H-1,3-benzodiazol-2(3H)-one (I and II) exhibit identical bond distances and angles except for the C—N—C—C torsion angle between the benzimidazolone backbone and the phenyl substituent, which has an effect on the crystal packing and supramolecular features. The structure of I contains a stronger C=O⋯H—N hydrogen-bonding interaction and a weaker π–π interaction between adjacent bezimidazolone moieties in comparison to II.The two polymorphic structures of 3-phenyl-1 I and II) of 3-phenyl-1H-1,3-benzodiazol-2(3H)-one, C13H10N2O, acquired from pentane diffusion into the solution in THF, are reported. The structures show negligible differences in bond distances and angles, but the C—N—C—C torsion angles between the backbone and the phenyl substituent, 123.02 (15)° for I and 137.18 (11)° for II, are different. Compound I features a stronger C=O⋯H—N hydrogen bond than that in II, while the structure of II exhibits a stronger π–π interaction than in I, as confirmed by the shorter intercentroid distance [3.3257 (8) Å in II in comparison to 3.6862 (7) Å in I]. Overall, the supramolecular interactions of I and II are distinct, presumably originating from the variation in the dihedral angle.The polymorphic structures ( The compound was prepared following the reported procedure using 1,1′-carbonyldiimidazole and N1-phenylbenzene-1,2-diamine in CH2Cl2  and blocks (II).Here we report two polymorphic structures of 3-phenyl-12.I) and blocks (II) in space groups C2/c and Pbca, respectively. The two polymorphic structures exhibit identical bond distances and angles, except for the dihedral angle of the phenyl substituent  and −137.18 (12)° for I and II, respectively. No additional differences are observed from an analysis of bond distances and angles.The title compounds crystallized as colorless needles  Å, while the corresponding distance in I is more elongated at 3.6862 (7) Å.Initial investigations of supramolecular features for y Figs. 2 and 3 ▸.I indicate the intermolecular interactions of C4⋯C2/C2⋯C4, C3A⋯C3A and C7—H7/H7—C7, whereas those in II correspond to C2⋯C5/C5⋯C2, C4—H4⋯C12/ C12⋯H4—C4, C7A⋯H6—C6/C6—H6⋯C7A, C3A⋯H6—C6/C6-H6⋯C3A and C3A⋯C6/C6⋯C3A contacts. The largest contributions to the Hirshfeld surface of I arises from H⋯H (44.4%), C⋯H/H⋯C (31.9%), and O⋯H/H⋯O (13.5%) contacts, whereas the contributions for II are H⋯H (45.8%), C⋯H/H⋯C (27.5%) and O⋯H/H⋯O (15.5%). Minor contributions include N⋯H/H⋯N (3.6%), C⋯C (3.2%), C⋯N/N⋯C (2.1%), C⋯O/O⋯C (1.4%) for I and C⋯C (5.4%), C⋯N/N⋯C (3.4%), N⋯H/H⋯N (3.2%), C⋯O/O⋯C (0.2%) for II.Minor intermolecular interactions are observed as faint red spots on the surface. The spots in 4.et al., 2016et al., 2013tert-butyl  chains. The distances between a nitrogen donor and an oxygen acceptor range from 2.79–2.84 Å, comparable to the values for I and II of 2.7786 (14) and 2.8453 (14) Å, respectively.A search for the title compound in the Cambridge Structural Database -one was prepared following a reported procedure  was stirred at room temperature overnight. The resulting white precipitate was filtered. An additional white precipitate was acquired by adding Et2O (10 mL) into the filtrate. Combined yield: 0.30 g (46%). 1H NMR : δ 10.75 , 7.58 , 7.45 , 7.17 , 7.10 , 7.06 . Pentane vapor diffusion into a solution of the compound in THF formed colorless needles and blocks.3-Phenyl-16.Crystal data, data collection, and refinement statistics are summarized in Table 310.1107/S2056989023003961/vm2281sup1.cifCrystal structure: contains datablock(s) global, I, II. DOI: 10.1107/S2056989023003961/vm2281Isup2.hklStructure factors: contains datablock(s) I. DOI: 10.1107/S2056989023003961/vm2281IIsup3.hklStructure factors: contains datablock(s) II. DOI: Click here for additional data file.10.1107/S2056989023003961/vm2281Isup4.cmlSupporting information file. DOI: 2260424, 2260423CCDC references:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


Reducing the necessary time to restore primary teeth improves the cooperation of paediatric patients. This study aimed to investigate the marginal integrity of restorations prepared with a bulk-fill resin-based composite (RBC) containing additional fragmentation chain transfer (AFCT) compared to a conventional RBC when light cured with a rapid high-irradiance (3 s) and a regular (10 s) curing mode.2 or high-irradiance: 3 s @ 3000 mW/cm2) and the used restorative material . After thermo-mechanical loading, the marginal integrity was analysed using scanning electron microscopy. A beta regression model and pairwise comparisons were used to statistically analyse the data.Forty class-II cavities were prepared in 40 primary molars. The molars were randomly divided into four groups based on the applied light-curing modes (regular: 10 s @ 1200 mW/cmp ≤ 0.05). The difference between the light-curing modes for the same RBC was not statistically significant (p ˃ 0.5).The mean marginal integrity (% ± SD) of the restorations for each group was as follows: Power Fill (10 s: 79.7 ± 15.6) (3 s: 77.6 ± 11.3), Prime (10 s: 69.7 ± 11.1) (3 s: 75.0 ± 9.7). The difference between the RBCs for the same light-curing mode was statistically significant (AFCT-containing bulk-fill RBC “Power Fill” achieves similar marginal integrity when light-cured with either high-irradiance or regular light-curing modes. “Power Fill” achieves better marginal integrity than the conventional RBC “Prime” regardless of the applied light-curing mode. Resin-based composites have become a solid restorative material option in both permanent and primary teeth –3. Some 2), and the incorporation of an β-allyl sulfone addition fragmentation chain transfer (AFCT) reagent in the matrix of the bulk-fill composite [Two major developments led to the possibility of such short polymerisation time: the introduction of high-power light emitting diode (LED) polymerisation units that can produce high radiant exitance  and the authorisation from the ethics committee was waived . Roots of the extracted molars were embedded in acrylic resin  3 mm below the cemento-enamel junction and mounted on custom-made holders. One standardised mesial or distal proximal cavity  was prepared in each primary molar. Proximal cavities were prepared with 80-µm cylindrical burs  rotating at 40,000 rpm in a high-speed contra-angle handpiece . The bur was exchanged after preparing four cavities. The molars were then randomised into four groups  based on the RBC they would be restored with and the light-curing mode they would be subjected to. The sample size was determined based on recent similar research [2 . The cavities were restored based on their experimental group as shown in Fig. 2, curing time = 10 s). In groups 3 and 4, each layer of both materials was subjected to high-irradiance light-curing . The composition of the used RBCs is shown in Table The molars were mounted and restored in a custom-made adjacent-tooth simulator with two plastic molars on each side on a laboratory desk. A stainless-steel matrix band  was inserted and fixed with a wooden wedge placed 1 mm below the gingival margin of the cavity. A universal adhesive  was scrubbed for 20 s on all cavity walls in self-etch mode, thinned with a gentle blow of air in order to evaporate the solvent, and then light-cured for 10 s at 1200 mW/cmAfter the storage time, impressions were taken for each restoration with an A-silicon material . The impressions were poured out , fixed on aluminium holders  and the formed replicas were sputter-coated with gold . After sputter coating, each replica was examined under a stereo microscope . The cemento-enamel junction buccally and lingually from the restoration was accentuated inside the epoxy resin using a fine scalpel. The replicas were quantitatively analysed for marginal integrity using scanning electron microscopy (SEM) at 20 kV and 200 × magnification . The marginal integrity for each restoration was expressed as a percentage of continuous margins in relation to the entire length of assessable margins. After this initial marginal analysis, all restorations underwent a thermo-mechanical loading (TML) inside an electronic masticator . The restorations were loaded on their occlusal part using a metal ball (diameter = 3 mm) for 400 000 loading cycles at 49 N . SimultaA-silicon impressions were taken again after TML and a final quantitative margin analysis using the same aforementioned protocol was carried out. The margin analysis was conducted by one calibrated and experienced operator (MZ) who was blinded to the groups and had only access to the SEM images. Figure 2 and 3 s @ 3000 mW/cm2). A beta regression model was set to analyse the data. Post-hoc pairwise comparisons between the groups of the same material with different light-curing modes “Power Fill (10 s)” vs. “Power Fill (3 s)” and “Prime (10 s)” vs. “Prime (3 s)” and between the groups of the same light-curing mode with different materials “Power Fill (10 s)” vs. “Prime (10 s)” and “Power Fill (3 s)” vs. “Prime (3 s)” were computed using the emmeans package [www.R-project.org).The marginal integrity  was set as the target variable. This variable was analysed with respect to the restoration type  and the light-curing modes (10 s @ 1200 mW/cm package . The sigp ˃ 0.1).Before TML, the achieved marginal integrity (% ± standard deviation SD) of each tested RBC  with each tested light-curing mode (regular “10 s” and high-irradiance “3 s”) was as follows: Power Fill (10 s: 92.1 ± 10.2) (3 s: 90.1 ± 5.4), Prime (10 s: 90.1 ± 6.9) (3 s: 90.3 ± 7.3). The difference between “Power Fill (10 s)” vs. “Power Fill (3 s)” and “Prime (10 s)” vs. “Prime (3 s)” and between “Power Fill (10 s)” vs. “Prime (10 s)” and “Power Fill (3 s)” vs. “Prime (3 s)” was not statistically significant (p ≤ 0.05). The difference between “Power Fill (10 s)” vs. “Power Fill (3 s)”, and between “Prime (10 s)” vs. “Prime (3 s)” was not statistically significant (p ˃ 0.5). Figure After TML, the achieved marginal integrity (% ± SD) of each tested RBC with each tested light-curing mode was as follows: Power Fill (10 s: 79.7 ± 15.6) (3 s: 77.6 ± 11.3), Prime (10 s: 69.7 ± 11.1) (3 s: 75.0 ± 9.7). The difference between “Power Fill (10 s)” vs. “Prime (10 s)”, and between “Power Fill (3 s)” vs. “Prime (3 s)” was statistically significant (p ˃ 0.1).Before TML, the achieved marginal integrity (% ± SD) of each tested RBC with each tested light-curing mode was as follows: Power Fill (10 s: 93.3 ± 10.6) (3 s: 89.5 ± 5.3), Prime (10 s: 89.3 ± 8.3) (3 s: 91.3 ± 6.7). The difference between “Power Fill (10 s)” vs. “Power Fill (3 s)” and “Prime (10 s)” vs. “Prime (3 s)”, and between “Power Fill (10 s)” vs. “Prime (10 s)” and “Power Fill (3 s)” vs. “Prime (3 s)” was not statistically significant (p ˃ 0.1). Figure After TML, the achieved marginal integrity (% ± SD) of each tested RBC with each tested light-curing mode was as follows: Power Fill 10 s: 83.4 ± 16.1) (3 s: 77.0 ± 11.9), Prime (10 s: 75.8 ± 14.1) (3 s: 76.9 ± 9.4). The difference between “Power Fill (10 s)” vs. “Power Fill (3 s)” and “Prime (10 s)” vs. “Prime (3 s)”, and between “Power Fill (10 s)” vs. “Prime (10 s)” and “Power Fill (3 s)” vs. “Prime (3 s)” was not statistically significant ( ± 16.1 (p ˃ 0.5).Before TML, the achieved marginal integrity (% ± SD) of each tested RBC with each tested light-curing mode was as follows: Power Fill (10 s: 88.9 ± 14.2) (3 s: 91.3 ± 7.0), Prime (10 s: 92.0 ± 4.7) (3 s: 88.2 ± 13.6). The difference between “Power Fill (10 s)” vs. “Power Fill (3 s)” and “Prime (10 s)” vs. “Prime (3 s)”, and between “Power Fill (10 s)” vs. “Prime (10 s)” and “Power Fill (3 s)” vs. “Prime (3 s)” was not statistically significant (p ≤ 0.05). The difference between “Power Fill (10 s)” vs. “Power Fill (3 s)”, and between “Prime (10 s)” vs. “Prime (3 s)” was also statistically significant (p ≤ 0.05). Figure After TML, the achieved marginal integrity (% ± SD) of each tested RBC with each tested light-curing mode was as follows: Power Fill (10 s: 71.0 ± 18.1) (3 s: 78.8 ± 14.0), Prime (10 s: 57.7 ± 15.4) (3 s: 71.2 ± 15.0). The difference between “Power Fill (10 s)” vs. “Prime (10 s)”, and between “Power Fill (3 s)” vs. “Prime (3 s)” was statistically significant (2) on the marginal integrity of an AFCT-containing bulk-fill RBC  and a conventional RBC in standardised deep class-II cavities in primary molars. The marginal integrity in this study was not affected by the different light-curing modes (the first null-hypothesis cannot be rejected), but indeed affected by the tested RBC (the second null-hypothesis must be rejected).This in-vitro study investigated the effect of a high-irradiance light curing  in the present study. The first attempts to evaluate dental restorative materials with regard to their marginal integrity date back to the 1970s . Some stThe here used loading force and frequency  is reported to be most frequently used to simulate the mechanical degradation of resin composite restorations in vitro . HoweverTaking the margins of the whole restoration (enamel and dentine) into consideration, employing the high-irradiance light-curing mode resulted in similar marginal integrities as the regular 10-s light-curing mode, regardless of the tested RBC. As for Power Fill, this result could be attributed to the presence of the AFCT reagent, which allows a step-like progression of the polymer chain and enhances the thermal and mechanical properties of RBCs, and to the fact that this RBC was actually developed to be compatible with high-irradiance light curing . Par et The bulk-fill RBC “Power Fill” showed better marginal integrity than the conventional RBC “Prime” regardless of the used light-curing mode. This is in contrast to the results of a recent study, which compared the same RBCs and concluded that both achieve similar marginal integrity in primary molars . This co2) does not actually help explain the observed similar marginal integrity. Nevertheless, Steffen et al. [2) or high-irradiance (3 s @ 2850 mW/cm2) modes. Similar findings (regarding shrinkage stress and degree of conversion) for AFCT-free RBCs were also reported by Par et al. [The conventional RBC “Prime” was not especially developed for high-irradiance light curing and does not contain an AFCT reagent. This makes the observed similar marginal integrity between both applied light-curing modes rather interesting. The fact that the radiant exposure in case of the high-irradiance light-curing mode is less than the regular light-curing mode  or regular light-curing modes (10 s @ 1200 mW/cm2).AFCT-containing high-viscous bulk-fill RBC “Power Fill” achieves similar performance with regard to marginal integrity of deep class-II cavities in primary molars when light-cured with high-irradiance (3 s @ 3000 mW/cmAFCT-containing high-viscous bulk-fill RBC “Power Fill” achieves better performance than the conventional RBC “Prime” with regard to marginal integrity in deep class-II cavities in primary molars regardless of the used light-curing mode.


Plasmodium falciparum, is driven by specific transcriptional programs, but it is unclear how most genes are activated or silenced at specific times. There is an association between transcription and spatial organization; however, the molecular mechanisms behind genome organization are unclear. While P. falciparum lacks key genome‐organizing proteins found in metazoans, it has all core components of the cohesin complex. To investigate the role of cohesin in P. falciparum, we functionally characterize the cohesin subunit Structural Maintenance of Chromosomes protein 3 (SMC3). SMC3 knockdown during early stages of the intraerythrocytic developmental cycle (IDC) upregulates a subset of genes involved in erythrocyte egress and invasion, which are normally expressed at later stages. ChIP‐seq analyses reveal that during the IDC, SMC3 enrichment at the promoter regions of these genes inversely correlates with gene expression and chromatin accessibility. These data suggest that SMC3 binding contributes to the repression of specific genes until their appropriate time of expression, revealing a new mode of stage‐specific gene repression in P. falciparum.The complex life cycle of the human malaria parasite,  Plasmodium falciparum, the cohesin subunit SMC3 plays a role in stage‐specific gene repression that is distinct from heterochromatin‐mediated repression of genes involved in antigenic variation and gametocytogenesis.In the human malaria parasite  Plasmodium falciparum, has a significant impact on human health in endemic regions  was found to play a role in the nuclear organization of centromeres, and knockdown led to defects in var gene transcription —key genome organizing proteins in metazoans , SMC3 (PF3D7_0414000), and a protein with the N‐terminal Rad21/Rec8 domain  is present in the P. falciparum cohesin complex  and stag , 24 (trophozoite), and 36 (schizont) hours post invasion (hpi). Using the macs2 peak calling algorithm  between significant peaks called in both clones for each time point, and we used this cohort of shared peaks for further analysis  of SMC3‐3HA‐ Dataset . No signet al, et al , “cell–cell adhesion” , “response to host” , and “antigenic variation” , “movement in host environment” (q = 3.3 × 10−3), “exit from host” (q = 0.026), and “entry into host” (q = 0.05). These categories include genes that are involved in invasion of or egress from the red blood cell such as ralp1 , as knockdown at the protein level could not be achieved after a single cell cycle , 69% at 24 hpi (Dataset q = 1.3 × 10−39), and 48% at 36 hpi  and ribonucleoprotein complex biogenesis (1.82 × 10−15), subunit organization (2.47 × 10−13), and assembly , glideosome‐associated protein 45 , merozoite surface protein 1 , and merozoite surface protein 9  does not inhibit parasite growth agrees with reports in S. cerevisiae and D. melanogaster in which normal growth and sister chromatid cohesion were achieved despite an 87 and 80% decrease, respectively, in Rad21, a core component of the cohesin complex  datasets do not provide evidence of typical enhancer‐promoter interactions found in other eukaryotes  is involved in transcription of invasion‐related genes via binding to the GTGCA motif, likely by interaction with the bromodomain protein 1  lysis during ring stage followed by a plasmagel  enrichment for late blood stages 24 h later. Another sorbitol treatment 6 h afterward places the 0 h time point 3 h after the plasmagel enrichment. Thus, the window of synchronicity for cultures is ± 3 h. Parasite development was monitored by Giemsa staining. Parasites were harvested at 1–5% parasitemia.Blood stage 3D7 io et al . Brieflyglms strain was generated using a two‐plasmid system (pUF1 and pL7) based on the CRISPR/Cas9 system previously described in Ghorbal et al (PfSMC3‐3HA‐glmS containing the single guide RNA (sgRNA) ‐encoding sequence 5′‐CCTAGAAAATTAGAACAATT‐3′ targeting the 3′ UTR of PF3D7_0414000. The pL7‐PfSMC3‐3HA‐glmS plasmid also contained the homology repair construct 5′‐AGATAGAGAGAGTTATATATCTAAAGGAACAAAGAATGAGGCCTACGAAATTATTAGCATTGTATAAAAAAAAAAAGAAAAAAAAAAGAAAAAAAAAAAAGATTATATATATAATATATGTTGACAATTAATAAATATATTTGTATATATCTGTTAACTAATTATGAAAATTTTTGAATCAATAAATTTTTTAAATAACAAAAAAAAAAAAAAATATATATATTATATATATATTTTATATTTTATATTTTCTTGTAATTTTTGTTTTTTTAGGAGGAAAAACATGCCCTAGAAAATggcggtggaTACCCTTACGATGTGCCTGATTACGCGTAtCCcTAtGAcGTaCCaGAcTAtGCGTACCCtTAtGAcGTtCCgGATTAtGCtcacggggtgTAAGCGGCCGCGGTCTTGTTCTTATTTTCTCAATAGGAAAAGAAGACGGGATTATTGCTTTACCTATAATTATAGCGCCCGAACTAAGCGCCCGGAAAAAGGCTTAGTTGACGAGGATGGAGGTTATCGAATTTTCGGCGGATGCCTCCCGGCTGAGTGTGCAGATCACAGCCGTAAGGATTTCTTCAAACCAAGGGGGTGACTCCTTGAACAAAGAGAAATCACATGATCTTCCAAAAAACATGTAGGAGGGGACAACAATTTGGTTTTGTTTTTTTCTTTAGGTTTTGAGAAAAACAAATAGGAAATACAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAATGTATTTTTACATATGCACTTGGATTATTTTATTTTTATTATTTTTCTTTATATAAAGTAAAAATATACATAAGTATGCTTATTTATTACATAAGAGTTTATTTAAGAAAGGTTTCTTTTTCATAATATTGTGTGCATGAGTTTTTTTTTATTTTATTTTTTTTTTTTATTTCTGTAACGAAAAGGATATTAAAAAAAATAATAAAA‐3′ . This homology repair construct comprises a 3 × Hemaglutinin (3HA) – encoding sequence followed by a glmS ribozyme‐encoding sequence  and 1.5 μM DSM1 (MR4/BEI Resources). Parasites reappeared approximately 3 weeks after transfection, and 5‐fluorocytosine was used to negatively select the pL7 plasmid.The SMC3‐3HA‐al et al . A 3D7 wet al (smc1 (PF3D7_1130700) and stag (PF3D7_1456500), respectively, were amplified using either SMC1‐HR‐FW/RV or STAG‐HR‐FW/RV (Dataset The SMC1‐3HA‐dd and STAG‐3HA‐dd strains were generated using the method of selection‐linked integration (SLI) previously described in Birnbaum et al . Homolog Dataset . The obt Dataset , followeE. coli (Agilent Technologies 200315). All the parasite lines were cloned by limiting dilution, and integration at the targeted genomic locus was confirmed by PCR (Dataset All cloning was performed using KAPA HiFi DNA Polymerase (Roche 07958846001), In‐Fusion HD Cloning Kit (Clontech 639649), and XL10‐Gold Ultracompetent  Dataset  and Sangglms clone  and WT culture , as a negative control, were synchronized. Late stage parasites (1.5 × 109 parasites) were enriched using Percoll density gradient separation and then cross‐linked with 1 ml 0.5 mM dithiobissuccinimidyl propionate  in DPBS for 60 min at 37°C  containing protease and phosphatase inhibitor cocktail (Thermo Fisher 78440) and 1 U/μl of Benzonase (Merck 71206). The lysates were cleared by centrifugation at 16,000 g for 10 min at 4°C. Supernatants were incubated with 25 μl of anti‐HA Dynabeads (Thermo Fisher 88836) overnight with rotation at 4°C. Beads were collected with a magnet and washed five times with 1 ml RIPA buffer, then five times with 1 ml DPBS, and then once with 1 ml 25 mM NH4HCO3 (Sigma 09830). The beads were reduced with 100 mM dithiothreitol (Sigma D9779), alkylated with 55 mM iodoacetamide (Sigma I1149), and subjected to on‐bead digestion using 1 μg of trypsin (Thermo Fisher 90059). The resulting peptides were desalted using C18 ziptips (Merck ZTC04S096) and sent for MS analysis.An SMC3‐3HA‐m/z, AGC 1e5  using an EASY‐Spray column, 50 cm × 75 μm ID, PepMap RSLC C18, 2 μm (Thermo Fisher ES803A) over a 70‐min gradient before nanoelectrospray using a Q Exactive HF‐X mass spectrometer (Thermo Fisher). The mass spectrometer was operated in a data‐dependent mode. The parameters for the full scan MS were as follows: resolution of 60,000 across 350–1,500 2, 1% IGEPAL CA‐630, and 1× protease inhibitor cocktail ) at 4°C and incubated on ice for 30 min. The cytoplasmic lysate was cleared with centrifugation . The pellet (containing the nuclei) was resuspended in 3.3 times less volume of nuclear extraction buffer  than cytoplasmic lysis buffer at 4°C, transferred to 1.5 ml sonication tubes , and sonicated for 5 min total (10 cycles of 30 s on/off) in a Diagenode Pico Bioruptor at 4°C. This nuclear lysate was cleared with centrifugation . Protein samples were supplemented with NuPage Sample Buffer (Thermo Fisher NP0008) and NuPage Reducing Agent (Thermo Fisher NP0004) and denatured for 10 min at 70°C. Proteins were separated on a 4–12% Bis‐Tris NuPage gel (Thermo Fisher NP0321) and transferred to a PVDF membrane with a Trans‐Blot Turbo Transfer system (Bio‐Rad). The membrane was blocked for 1 h with 1% milk in PBST  at 25°C. HA‐tagged proteins and histone H3 were detected with anti‐HA  and anti‐H3  primary antibodies, respectively, followed by donkey anti‐rabbit secondary antibody conjugated to horseradish peroxidase . SMC3 was detected with an in‐house‐generated anti‐SMC3 antibody , followed by sheep HRP anti‐mouse secondary antibody . HRP signal was developed with SuperSignal West Pico Plus or Femto chemiluminescent substrate  and imaged with a ChemiDoc XRS+ (Bio‐Rad).Parasites were washed once with Dulbecco's phosphate‐buffered saline , then resuspended in cytoplasmic lysis buffer  was amplified from cDNA of 3D7 wild‐type parasites and inserted into the pGEX expression vector. The plasmid was verified by sequencing and used to transform SHuffle T7 expression bacteria (New England BioLabs C3026J). Recombinant protein expression was induced with 100 μM IPTG overnight with shaking at 16°C, and the pellet was frozen at −80°C until use. Soluble GST‐tagged recombinant protein was purified using Glutathione Sepharose 4 Fast Flow (Cytiva GE17‐5132‐03), according to manufacturer's instructions. Briefly, the bacterial pellet was resuspended in cold lysis buffer . The lysate was sonicated on ice with a probe sonicator . The sample was centrifuged at 18,000 g for 1 h at 4°C, and the soluble lysate was incubated with Glutathione Sepharose beads (Cytiva GE17‐5132‐03) overnight with gentle tumbling at 4°C. The next day, the lysate with beads was loaded onto chromatography columns , and the beads were washed with cold PBS containing 5 mM EDTA. Recombinant protein was eluted with elution buffer , buffer exchanged into PBS , and stored at −20°C until use. Final protein concentration was obtained using Protein Assay (BioRad).The coding sequence for the N‐terminal region (amino acids 1‐253) of th, 2013). All experiments were approved by the Ethics Committee CETEA Institute Pasteur and registered under the reference Permit Number dap180040, issued in 2018. The PfSMC3 antibody was purified from pooled mouse serum using rProtein G Agarose (Invitrogen 15920) according to manufacturer's instructions. The final purified antibody was dialyzed in PBS using a Float‐A‐Lyzer (Repligen). Purified antibodies were stored at −20°C in 50% Glycerol.Purified recombinant protein was used to immunize C57BL/6 mice. Ten micrograms were used for the first immunization in Freund's Complete Adjuvant (Sigma F5881) and three subsequent boosters in Freund's Incomplete Adjuvant (Sigma F5506). Animals were housed in the Institute Pasteur animal facilities, accredited by the French Ministry of Agriculture for performing experiments on live rodents. Work on animals was performed in compliance with French and European regulations on care and protection of laboratory animals . Parasite cultures were centrifuged at 800 g for 3 min at 25°C. Medium was removed and the RBCs were lysed with 10 ml 0.075% saponin (Sigma S7900) in DPBS at 37°C. The parasites were centrifuged at 3,250 g for 3 min at 25°C and washed twice with 10 ml DPBS at 4°C. The supernatant was removed, and the parasite pellet was resuspended in 1 ml 0.5 mM dithiobissuccinimidyl propionate  in DPBS for 30 min at 25°C  were washed twice with ChIP dilution buffer  using a DynaMag magnet (Thermo Fisher 12321D). The beads were resuspended in 200 μl 0.02% Tween‐20 in DPBS with 1 μg of anti‐HA antibody (Abcam ab9110) or IgG (Diagenode C15410206) and incubated with rotation at 4°C for 6 h.g and the supernatant was removed. The pellet was resuspended in 1 ml cytoplasmic wash buffer  and rotated for 10 min. The lysates were centrifuged for 5 min at 5,000 g, and the supernatant was removed. The pellet was resuspended in 200 μl nuclear lysis buffer , transferred to 1.5 ml sonication tubes (Diagenode C30010016), and sonicated for 2.5 min total (5 cycles of 30 s on/off) in a Diagenode Pico Bioruptor. The sonicated extracts were centrifuged at 13,500 g for 10 min. The supernatant was diluted with 400 μl dilution buffer 2 .All further steps were performed at 4°C. The cross‐linked parasites were resuspended in 900 μl of cytoplasmic lysis buffer  and rotated for 30 min. The lysates were centrifuged for 5 min at 2,000 PfSMC3 antibody (1:500 in 1% milk‐PBST) followed by sheep HRP anti‐mouse secondary antibody . Histone H3 was detected with Anti‐H3 , followed by donkey HRP anti‐rabbit secondary antibody . HRP signal was developed with SuperSignal West Pico Plus or West Femto chemiluminescent substrate  and imaged with a ChemiDoc XRS+ (Bio‐Rad).Using a DynaMag magnet, the antibody‐conjugated Dynabeads were washed once with 200 μl of dilution buffer 1  and resuspended in 200 μl dilution buffer 2 . The washed antibody‐conjugated Dynabeads were added to the diluted nuclear lysate and incubated overnight with rotation at 4°C. The beads were collected on a DynaMag magnet and the supernatant was removed and kept as flow through. The beads were washed three times with 200 μl of wash buffer . The beads were resuspended in 30 μl of 2× NuPage Sample Buffer (Thermo Fisher NP0008) and 3.4 μl of NuPage Reducing Agent (Thermo Fisher NP0004) and denatured for 5 min at 95°C. The eluate was separated from the beads with the DynaMag magnet. Lysates and the bead eluate were separated on a 3‐8% Tris‐Acetate NuPage gel (Thermo Fisher EA0375) and transferred to a PVDF membrane with a Trans‐Blot Turbo Transfer system (Bio‐Rad). Membranes were blocked with 1% milk‐PBST . HA‐tagged SMC1 and STAG proteins were detected with anti‐HA conjugated to HRP . SMC3 was detected with anti‐et al, et al  in PBS for 30 min. Cells were incubated with anti‐HA  and/or anti‐HP1 [from  with DAPI  for 45 min followed by three 10 min washes with 0.5% Tween®20/PBS. Cells were washed once more with PBS and placed onto polyethyleneimine‐coated slides (Thermo Scientific 30‐42H‐RED‐CE24). Once adhered to the slide, cells were washed twice with PBS and mounted with VectaShield® (Vector Laboratories). Images were acquired using a Deltavision Elite imaging system , and Fiji (http://fiji.sc) was used for analysis using the least manipulation possible.iRBCs were washed once with DPBS (Thermo Fisher 14190) at 37°C and fixed in suspension in 4% paraformaldehyde (EMS 15714) with 0.0075% glutaraldehyde (EMS 16220) in PBS for 20 min at 25°C, as described previously , 24  and 36 hpi . Parasite culture was centrifuged at 800 g for 3 min at 25°C. Medium was removed and the RBCs were lysed with 10 ml 0.075% saponin (Sigma S7900) in DPBS at 37°C. The parasites were centrifuged at 3,250 g for 3 min at 25°C and washed with 10 ml DPBS at 37°C. The supernatant was removed, and the parasite pellet was resuspended in 10 ml of PBS at 25°C. The parasites were cross‐linked by adding methanol‐free formaldehyde  and incubating with gentle agitation for 10 min at 25°C. The cross‐linking reaction was quenched by adding glycine  and incubating with gentle agitation for 5 min at 25°C. Parasites were centrifuged at 3,250 g for 5 min at 4°C and the supernatant removed. The pellet was washed with DPBS and centrifuged at 3,250 g for 5 min at 4°C. The supernatant was removed, and the cross‐linked parasite pellets were snap‐frozen.Two clonal populations (clone A and B) of SMC3‐3HA‐For each time‐point, 200 μl of Protein G Dynabeads (Invitrogen 10004D) were washed twice with 1 ml ChIP dilution buffer  using a DynaMag magnet (Thermo Fisher 12321D). The beads were resuspended in 1 ml ChIP dilution buffer with 8 μg of anti‐HA antibody (Abcam ab9110) and incubated on a rotator at 4°C for 6 h.g at 4°C, the supernatant was removed, and the pellet was resuspended in SDS lysis buffer  at 4°C (3.6 ml for the 12 hpi sample and 1.8 ml for the 24 ad 36 hpi samples). The liquid was distributed into 1.5 ml sonication tubes  and sonicated for 12 min total (24 cycles of 30 s on/off) in a Diagenode Pico Bioruptor at 4°C. The sonicated extracts were centrifuged at 13,500 g for 10 min at 4°C and the supernatant, corresponding to the chromatin fraction, was kept. The DNA concentration for each time point was determined using the Qubit dsDNA High Sensitivity Assay Kit (Thermo Fisher Scientific Q32851) with a Qubit 3.0 Fluorometer (Thermo Fisher Scientific). For each time point, chromatin lysate corresponding to 100 ng of DNA was diluted in SDS lysis buffer  and kept as “input” at −20°C. For clones A and B, chromatin lysate corresponding to 19 and 18 μg (12 hpi), 2 and 1.4 μg (24 hpi) and 3 and 5.2 μg (36 hpi) of DNA, respectively, was diluted 1:10 in ChIP dilution buffer at 4°C.The cross‐linked parasites were resuspended in 4 ml of lysis buffer  at 4°C, and 10% Nonidet‐P40 was added . The parasites were lysed in a prechilled dounce homogenizer (200 strokes for 12 hpi parasites and 100 strokes for 24 and 36 hpi parasites). The lysates were centrifuged for 10 min at 13,500 Low salt wash buffer  at 4°C.High salt wash buffer  at 4°C.LiCl wash buffer  at 4°C.TE wash buffer  at 25°C.Using a DynaMag magnet, the antibody‐conjugated Dynabeads were washed twice with 1 ml ChIP dilution buffer and resuspend in 100 μl of ChIP dilution buffer at 4°C. Then the washed antibody‐conjugated Dynabeads were added to the diluted chromatin sample and incubated overnight with rotation at 4°C. The beads were collected on a DynaMag into eight different tubes per sample, the supernatant was removed, and the beads in each tube were washed for 5 min with gentle rotation with 1 ml of the following buffers, sequentially:After the washes, the beads were collected on a DynaMag, the supernatant was removed, and the beads for each time point were resuspended in 800 μl of elution buffer and incubated at 65°C for 30 min with agitation . The beads were collected on a DynaMag and the eluate, corresponding to the “ChIP” samples, was transferred to a different tube.g at 4°C to separate phases. The aqueous top layer was transferred to another tube and mixed with 30 μg glycogen (Thermo Fisher 10814) and 5 M NaCl . Eight hundred microliters 100% EtOH at 4°C were added to each sample, which was then incubated at −20°C for 30 min. The DNA was pelleted by centrifugation for 10 min at 13,500 g at 4°C, washed with 500 μl 80% EtOH at 4°C, and centrifuged for 5 min at 13,500 g at 4°C. After removing the EtOH, the pellet was dried at 25°C and all DNA for each sample was resuspended in 30 μl 10 mM Tris–HCl, pH 8 total. The DNA concentration and average size of the sonicated fragments was determined using a DNA high sensitivity kit (Agilent 5067‐4626) and the Agilent 2100 Bioanalyzer. Libraries for Illumina Next Generation Sequencing were prepared with the MicroPlex library preparation kit (Diagenode C05010014), with KAPA HiFi polymerase (KAPA biosystems) substituted for the PCR amplification. Libraries were sequenced on the NextSeq 500 platform (Illumina).For purification of the DNA, both “ChIP” and “Input” samples were incubated for approximately 10 h at 65°C to reverse the crosslinking. Two hundred microliters of TE buffer followed by 8 μl of RNaseA  were added to each sample, which was then incubated for 2 h at 37°C. Four microliters Proteinase K  were added to each sample, which was then incubated for 2 h at 55°C. Four hundered microliters phenol:chloroform:isoamyl alcohol  were added to each sample, which was then mixed with vortexing and centrifuged for 10 min at 13,500 P. falciparum genome  were mapped to the P. falciparum reference genome feature file  was added to one half of the culture for two rounds of parasite replication (approximately 96 h). Parasites were then re‐synchronized and three technical replicates (with and without glucosamine) were harvested at 12, 24, and 36 hpi. RBCs were lysed in 0.075% saponin (Sigma S7900) in PBS at 37°C, centrifuged at 3,250 g for 5 min, washed in PBS, centrifuged at 3,250 g for 5 min, and resuspended in 700 μl QIAzol reagent (Qiagen 79306). RNA was extracted using an miRNeasy Mini kit (Qiagen 1038703) with the recommended on‐column DNase treatment. Total RNA was poly (A) selected using the Dynabeads mRNA Purification Kit (Thermo Fischer Scientific 61006). Library preparation was performed with the NEBNext® Ultra™ II Directional RNA Library Prep Kit for Illumina® (New England Biolabs E7760S) and paired end sequencing was performed on the Nextseq 550 platform (Illumina). Sequenced reads were mapped to the P. falciparum genome  using the built‐in tool at PlasmoDB.org .A WT and SMC3‐3HA‐et al, et al, et al  of each sample to the microarray data from  was added to one half for approximately 96 h before starting the growth curve. The parasites were tightly re‐synchronized and diluted to ~0.2% parasitemia (5% hematocrit) at ring stage. The growth curve was performed in a 96‐well plate (200 μl culture per well) with three technical replicates per condition per blood. Every 24 h, 5 μl of the culture were fixed in 45 μl of 0.025% glutaraldehyde in PBS for 1 h at 4°C. After centrifuging at 800 g for 5 min, free aldehyde groups were quenched by re‐suspending the iRBC pellet in 200 μl of 15 mM NH4Cl in PBS. A 1:10 dilution of the quenched iRBC suspension was incubated with Sybr Green I (Sigma S9430) to stain the parasite nuclei. Quantification of the iRBCs was performed in a CytoFLEX S cytometer (Beckman Coulter), and analysis with was done with FlowJo Software. Growth rate was calculated and expressed as the parasitemia at Day X divided by the starting average parasitemia (Day 1) of the correspondent culture.Parasite growth was measured as described previously (Vembar BioRender.com.The synopsis image for this manuscript was created with Catarina Rosa: Conceptualization; investigation; visualization; writing – original draft; writing – review and editing. Parul Singh: Conceptualization; data curation; formal analysis; validation; visualization; writing – original draft; writing – review and editing. Patty Chen: Investigation; visualization. Ameya Sinha: Investigation. Aurélie Claës: Investigation. Peter R Preiser: Supervision; funding acquisition. Peter C Dedon: Supervision; funding acquisition. Sebastian Baumgarten: Conceptualization; formal analysis; supervision; validation; visualization; writing – original draft; writing – review and editing. Artur Scherf: Supervision; funding acquisition. Jessica M Bryant: Conceptualization; formal analysis; supervision; funding acquisition; visualization; writing – original draft; project administration; writing – review and editing.The authors declare that they have no conflict of interest.Expanded View Figures PDFClick here for additional data file.Dataset EV1Click here for additional data file.Dataset EV2Click here for additional data file.Dataset EV3Click here for additional data file.Dataset EV4Click here for additional data file.Dataset EV5Click here for additional data file.Dataset EV6Click here for additional data file.Dataset EV7Click here for additional data file.Dataset EV8Click here for additional data file.Dataset EV9Click here for additional data file.Dataset EV10Click here for additional data file.Dataset EV11Click here for additional data file.Dataset EV12Click here for additional data file.Dataset EV13Click here for additional data file.Dataset EV14Click here for additional data file.Dataset EV15Click here for additional data file.Dataset EV16Click here for additional data file.Dataset EV17Click here for additional data file.Dataset EV18Click here for additional data file.Dataset EV19Click here for additional data file.Dataset EV20Click here for additional data file.Dataset EV21Click here for additional data file.Dataset EV22Click here for additional data file.PDF+Click here for additional data file.Source Data for Figure 1Click here for additional data file.Source Data for Figure 2Click here for additional data file.Source Data for Figure 4Click here for additional data file.


The crystal structures of a dihydrofurylsilane and a dihydrofurylgermane are reported. Hirshfeld surface analyses were performed to investigate the intermolecular interactions. 4H5O)4 (1) and Ge(C4H5O)4 (2) are dihydrofuryl compounds of silicon and germanium and are useful building blocks for the functionalization of these elements. Both structures crystallize in space group P21/n in the monoclinic crystal system with two molecules in the asymmetric unit: the Si and Ge atoms adopt slightly distorted tetrahedral geometries, while the C4H5O moieties exhibit shallow envelope conformations. Through a Hirshfeld surface analysis of the structures, interactions within the crystal packing could be elucidated: compound 1 features a polymeric chain in the (101) plane via C—H⋯O hydrogen bonds whereas in 2 C—H⋯O hydrogen bonds create a polymeric chain in the (010) plane.The title compounds Si(C The two compounds are already known in the literature  were prepared by Lukevits and co-workers in the 1980s  −x, −y, −z] with a distance of 2.22 (4) Å was identified by the red spots on the Hirshfeld surface. However, these red spots show only a small proportion of the interactions indicated in the fingerprint. Furthermore, a C30⋯H24Bii  van der Waals interaction with a separation of 2.796 (16) Å was also identified. The contribution of the C⋯H interactions is 10.7%, which is a low contribution to the crystal packing. Besides these interactions, H⋯O interactions could be identified and contribute 19.2% of the structure in the solid state. Hydrogen bonds between C—H⋯O, which are indicated by red spots on the Hirshfeld surface are listed in Table 5A⋯O5ii, C27—H27B⋯O3iii and C31—H31A⋯O4 can be described as having a D11(2) graph-set motif. C23—H23B⋯O5i is described by et al., 1990B⋯O3iii and C31—H31A⋯O4, a part of the crystal packing is defined along the [101] direction  Å and H7B⋯H7Bi  at 2.20 (4) Å are visible as red spots and could be identified as close interactions by the Hirshfeld surface. The contribution of the van der Waals interactions is slightly lower at 10.0%. The interaction C32⋯H4Aii  at 2.79 (2) Å could also be detected on the Hirshfeld surface. As in the case of 1, interactions of the form H⋯O could be determined, which contribute 18.3% to the crystal packing. These hydrogen bonds were detected by red spots on the Hirshfeld surface and are shown in Table 6A⋯O7i and C23–H23A⋯O4ii can be described by the graph-set motif D11(2). In contrast, the hydrogen bond C31—H31A⋯O7iii is described by the graph-set motif et al., 1990A⋯O7iii and C31—H31A⋯O7i, a part of the crystal packing, which forms a plane in the [010] direction, can be seen in Fig. 8For the Hirshfeld surface analysis of ed Fig. 6. The dised Fig. 6. The dis4.et al., 2016et al., 2022et al., 20201 and 2. They also display similar (DHF)C—Si—C(DHF) bond angles and also a slightly distorted tetrahedron. In addition, a deviation in the planarity of the dihydrofuryl rings was found there. An extended search for 3-silanes revealed the compounds (trimethyl)silane -5-methyl-4-(t-butyldiphenylsilyl)-2,3-dihydro-furan-2-carboxylic acid (1′-phenylethyl)amide -furanone silane was also found in the database when searching for (2-furanyl)silane germane and an extended search for 3-germane found no hits.5.1 and 2 have already been described by Lukevits and Ertschak , tert-butyllithium  was added at 228 K to a solution of 2,3-dihydrofuran  in diethyl ether (approx. 100 ml). The reaction solution was stirred for 1 h at room temperature. Then, tetrachlorosilane  was added at 243 K and the reaction solution was stirred for 1 h. The resulting solid was separated by inert filtration. The obtained solution was concentrated in vacuo and crystallized at 243 K. The solvent was removed and the solid was washed with cold diethyl ether. The product tetrakissilane (1)  was obtained as colourless blocks.Compound 1H NMR: : δ = 2.25 , 4.06 , 5.88  ppm. {1H}13C NMR: : δ = 31.4 , 71.0 , 117.8 , 155.1  ppm. {1H}29Si NMR: : δ = −51.40  ppm.2), tert-butyllithium  was added at 228 K to a solution of 2,3-dihydrofuran  in diethyl ether (approx. 100 ml). The reaction solution was stirred for 1 h at rt. Tetrachlorogermane  was added at 213 K and the reaction solution was stirred for 1 h. The resulting solid was separated by inert filtration. The obtained solution was concentrated in vacuo and crystallized at 243 K. The solvent was removed, and the solid was washed with cold diethyl ether. The product tetrakisgermane (2)  was obtained as colourless blocks.For the synthesis of tetrakisgermane : δ = 2.26 , 4.05 , 5.62  ppm. {1H}13C NMR: : δ = 30.8 , 71.0 , 113.8 , 155.7  ppm.6.A,B, H23A,B, H24A,B, H27A,B and H31A,B for compound 1 and all H atoms for compound 2 were refined independently. Other H atoms were positioned geometrically (C—H = 0.95–1.00 Å) and were refined using a riding model, with Uiso(H) = 1.2Ueq(C) for CH2 and CH hydrogen.Crystal data, data collection and structure refinement details are summarized in Table 710.1107/S2056989023003158/hb8058sup1.cifCrystal structure: contains datablock(s) 1. DOI: 10.1107/S2056989023003158/hb80581sup2.hklStructure factors: contains datablock(s) 1. DOI: Click here for additional data file.10.1107/S2056989023003158/hb80581sup4.cmlSupporting information file. DOI: 10.1107/S2056989023003158/hb80582sup3.hklStructure factors: contains datablock(s) 2. DOI: Click here for additional data file.10.1107/S2056989023003158/hb80582sup5.cmlSupporting information file. DOI: 2254180CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


The title compound exhibits I⋯π halogen bonding and π-stacking in its extended structure. 18H8I2, is an ethynyl-substituted anthracene. The C—C—I bond angles deviate from 180°, being affected by intermolecular I⋯π interactions. These interactions form a two-dimensional supramolecular structure further supported by offset π–π stacking of neighboring anthracene moieties.The title compound, C Two-dimensional crystals can have unique properties with applications in electronics, biomedicine, and sensors —I moieties and the π-electrons of the adjacent anthracene rings  [I1⋯Cg1 = 3.528 (4) Å and C16—I1⋯Cg1 = 151.2 (3)°] and I2 has a short contact to the centroid of the C9–C14 ring (Cg2) [I2⋯Cg2 = 3.767 (4) Å and C18—I2⋯Cg2 = 150.1 (3)°]. The bent nature of the C—I⋯centroid interactions leads to short I⋯C contacts ranging from 3.352 (4) to 3.655 (4) Å. The shorter contact between I1 and Cg1 appears to influence more significantly the bending of the entire alkynyl substituent [C1—C15—I1 = 173.8 (3)° versus C8—C17—I2 = 178.7 (3)°], notably pulling the I1 atom away from the central ring of the neighboring anthracene molecule and toward its C2–C7 centroid.The crystal stucture represents the first example of an ethynyl–anthracene halogenated with iodine Fig. 1. The C—Igs Fig. 2, where Iet al., 2008et al., 2014et al., 2015tert-butyl)[4-(iodoethynyl)phenyl]carbamate  plane Fig. 3. Intereet al., 2021N-iodosuccinimide  and AgNO3  were added to dry dimethylformamide (5 ml), and the resulting mixture was stirred under nitrogen. After 5 h, the reaction mixture was diluted with EtOAc (30 ml) and washed with H2O (5 × 30 ml). The organic layer was dried in vacuo, resulting in an orange solid. The product was crystallized from the orange solid using vapor–vapor diffusion (CH2Cl2/hexanes).The procedure was modeled after an analogous functionalization of an alkynylsilane : δ 8.33 , 7.82 .Crystal data, data collection, and structural refinement details are summarized in Table 110.1107/S2414314623005539/hb4433sup1.cifCrystal structure: contains datablock(s) I, global. DOI: 10.1107/S2414314623005539/hb4433Isup2.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S2414314623005539/hb4433Isup3.cmlSupporting information file. DOI: 2191397CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


E-configuration with respect to the double bond of the hydrazone bridge and with an acyl–hydrazone (—CH=N—NH—CO—) torsion angle of 166.0 (3)°. The molecule exhibits a non-planar conformation, likely induced by packing requirements.The title aroylhydrazone ether exists in an  22H19N3O4, shows a non-coplanar conformation, with dihedral angles between the phenyl rings of 73.3 (1) and 80.9 (1)°. These deformations are induced by the crystal packing that is mainly governed by N—H⋯O and C—H⋯O hydrogen bonds, forming a mono-periodic arrangement parallel to the b axis.The molecular structure of the title compound, C The active pharmacophore group, —CH=N—NH—C=O—, present in a hydrazone is primarily responsible for its broad spectrum of biological aspects  Å is slightly shorter than that of 1.397 (4) Å determined in the corresponding derivative having a thienyl ring replacing the p-nitrophenyl group , 3315 ν(N—H), 1606 ν(C=Nazomethine). LC–MS (FAB) m/z: [M + H]+ calculated for C22H19N3O4; 390.1446; found 390.1448.Yield: 0.29 g, 79%; melting point (m.p.): 531–533 K; FT–IR: 1636 ν I, global. DOI: 10.1107/S2056989023003948/wm5682Isup2.hklStructure factors: contains datablock(s) I. DOI: Click here for additional data file.10.1107/S2056989023003948/wm5682Isup3.cmlSupporting information file. DOI: 2232132CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


E)-1-(4-bromophenyl)-3-(2-methylphenyl)prop-2-en-1-one, the molecules are linked into chains by weak C—H⋯O interactions along the b axis. Successive chains form a zigzag structure along the c axis, and these chains are connected to each other by face-to-face π–π stacking interactions along the a axis, forming layers parallel to the (001) plane. The crystal structure maintains its stability via van der Waals interactions between the layers.In the crystal of °, and the configuration about the C=C bond is E. In the crystal, the molecules are linked into chains by weak C—H⋯O interactions along the b axis. Successive chains form a zigzag structure along the c axis, and these chains are connected to each other by face-to-face π–π stacking interactions along the a axis. These layers, parallel to the (001) plane, are linked by van der Waals interactions, thus consolidating the crystal structure. Hirshfeld surface analysis showed that the most significant contacts in the structure are H⋯H (43.1%), C⋯H/H⋯C (17.4%), Br⋯H/H⋯Br (14.9%), C⋯C (11.9%) and O⋯H/H⋯O (9.8%).In the title compound, C The molecule is approximately planar, as indicated by the torsion angles C10—C5—C4—C3 = 1.9 (5)°, C9—C4—C3—C2 = −4.4 (5)°, C4—C3—C2—C1 = −176.3 (3)°, C3—C2—C1—C11 = −168.2 (3)°, C2—C1—C11—C12 = 15.9 (4)° and Br1—C14—C15—C16 = 178.5 (2)°. The dihedral angle between the planes of the 2-methylphenyl and 4-bromophenyl rings is 23.49 (15)°.The title compound Fig. 1 is comp3.C(5) chains  Å, slippage = 1.890 Å; Cg2⋯Cg2a = 3.9420 (18) Å, slippage = 1.942 Å; symmetry code: (a) x − 1, y, z; Cg1 and Cg2 are the centroids of the 2-methylphenyl (C4—C9) and 4-bromophenyl (C11–C16) rings, respectively]. They form layers parallel to the (001) plane through van der Waals interactions, thus consolidating the crystal structure.In the crystal, the molecules are linked into is Fig. 3 and thesCrystalExplorer17.5 ; 43.1%], C⋯H/H⋯C , Br⋯H/H⋯Br , C⋯C  and O⋯H/H⋯O  interactions contribute the most to the surface contacts. The crystal packing is additionally influenced by Br⋯C/C⋯Br (2.0%), Br⋯Br (0.8%), N⋯N (2.6%) and O⋯C/C⋯O (0.2%) contacts. The Hirshfeld surface study confirms the significance of H-atom interactions in the packing formation. The large number of H⋯H, C⋯H/H⋯C, Br⋯H/H⋯Br, C⋯C and O⋯H/H⋯O interactions indicates that van der Waals interactions and hydrogen bonding are important in the crystal packing -1,3-diphenylprop-2-en-1-one’ unit in the Cambridge Structural Database  Å], forming layers lying parallel to the ab plane. In the crystal of RUCKIM, the molecules are linked through type II halogen bonds, forming a sheet structure parallel to the bc plane. Weak intermolecular C—H⋯π interactions are observed between the sheets. In the crystal of XOLLOC, molecules are linked via pairs of C—H⋯O interactions with an via C—H⋯π interactions. In the crystal of OBIYUW01, molecules are linked by C—H⋯π interactions between the bromophenyl and fluorophenyl rings, resulting in a two-dimensional layered structure parallel to the ab plane. The molecular packing is consolidated by weak Br⋯H and F⋯H contacts.In the crystal of KOCZUA, the shortest intermolecular contacts are Cl⋯O [3.173 (3) Å]; these link the molecules to form a 25.et al., 2021v/v) solution at room temperature.The title compound was synthesized using a reported procedure (Chithiraikumar 6.Uiso(H) = 1.2Ueq(C) for aromatic H atoms, and C—H = 0.98 Å and Uiso(H) = 1.5Ueq(C) for methyl H atoms.Crystal data, data collection and structure refinement details are summarized in Table 310.1107/S2056989023007387/tx2073sup1.cifCrystal structure: contains datablock(s) I, global. DOI: 10.1107/S2056989023007387/tx2073Isup2.hklStructure factors: contains datablock(s) I. DOI: 2290092CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


This salt consists of a complex [Ni(OPD)2(H2O)2]2+ cation with two bidentate OPD ligands and trans aqua ligands, and a non-coordinating NDS2– anion, which is the double-deprotonated form of H2NDS. The NiII atom is situated at a center of inversion and exhibits a slightly tetragonally distorted {O2N4} octahedral coordination environment, with four shorter equatorial Ni—N bonds [2.0775 (17) and 2.0924 (18) Å] and a longer axial Ni—O bond [2.1381 (17) Å]. The OPD ligand is located about an inversion center and is nearly coplanar with the NiN4 plane [dihedral angle 0.95 (9)°]. In the crystal, the cations and anions are connected by charge-assisted intermolecular N—H⋯O and O—H⋯O hydrogen-bonding interactions into the tri-periodic network structure. Hirshfeld surface analysis indicates that the most important contributions to the crystal packing are from H⋯H (44.1%), O⋯H/H⋯O (34.3%), C⋯H/H⋯C (14.8%) C⋯C (6.5%) (involving the cations) and O⋯H/H⋯O (50%), H⋯H (25%), C⋯H/H⋯C (15.3%), C⋯C (8.2%) (involving the anions) interactions.The reaction of  Its reactions with carboxylic acids and their derivatives produce the important class of benzimidazoles  or its deprotonated form (sulfonates) are of interest in supramolecular chemistry 2]·NDS, where the NDS2– anion is not part of the metal coordination sphere.In this work, we focus on the synthesis, crystal structure, and Hirshfeld surface analysis of a nickel(II) complex,  and a pair of longer axial Ni—O bonds [2.1381 (17) Å]. The OPD ligand, likewise located over a crystallographic inversion center at the middle of the central C11—C11 bond, is almost coplanar with the NiN4 plane, with a dihedral angle of 0.95 (9)°. The deviation of the ideal octahedral coordination sphere around nickel might be explained as follows: The inflexible nature of the OPDA ring system with an N⋯N distance between the amino groups of 2.770 (2) Å determines the N2—Ni1—N1 and N2—Ni1—N1 bite angles of 83.26 (7) and 96.74 (7) °, respectively.The structures of the molecular entities of the title compound are shown in Fig. 13.2(H2O)2]2+ cation and the NDS2– anion are associated via charge-assisted intermolecular O—H⋯O and N—H⋯O hydrogen bonds (Table 12(H2O)2]2+ cation forms N—H⋯O and O—H⋯O hydrogen bonds with six neighboring organic anions whereby the two aqua and two OPD ligands act solely as hydrogen-bonding donor groups . Additionally, cations and neighboring dianions are linked through O1W—H1WA⋯O3iv and N2—H2B⋯O3i hydrogen bonds. The molecules stack along [001], thereby forming a consolidated tri-periodic supramolecular network s Table 1. Each 2+ cation and the NDS2– dianion. The dnorm surface has twelve bright-red spots on the Hirshfeld surface for the cation and anion each 2]2+ cation and NDS2– anion, respectively; Fig. 6b and 6f). O⋯H/H⋯O and C⋯H/H⋯C, interactions in the cation, and H⋯H and C⋯H/H⋯C interactions in the dianion follow with contributions of 34.3, 14.8, 25 and 15.3%, respectively . Other minor contributions are from C⋯C (6.5%) and C⋯O (0.3%) contacts in the cation, and from C⋯C (8.2%), C⋯O (0.3%), O⋯O (0.1%) and S⋯H (0.1%) contacts in the dianion. The O⋯H/H⋯O contacts are visible as a spike with a sharp tip on the side of the corresponding two-dimensional fingerprint plot, which is indicative of strong intermolecular interactions between atoms. On the other hand, the C⋯H/H⋯C contacts form less pronounced spikes, suggesting that these interactions are much weaker.The supramolecular interactions discussed above were quantitatively investigated and visualized using Hirshfeld surface analysis performed with ch Fig. 5, resultily Fig. 6c,d,g,h.4.et al., 2016o-phenylenediamine moiety were identified. Out of these, 129 compounds were metal complexes, while 78 compounds were organic salts. One organic salt comprising protonated o-phenylenediamine and 1,5-naphthalenedisulfonate has been studied  sulfate heptahydrate  and disodium naphthalene-1,5-disulfonate  in 10 ml of the same mixed ethanol/water solvent. The resulting mixture was heated under reflux and stirred for 40 min. After 5 d of slow solvent evaporation at room temperature, a light-green crystalline product was obtained with a yield of 65% (based on Ni). Elemental analysis calculated (%) for C22H26N4NiO8S2: C 44.24, H 4.39, N 9.38; found: C 44.18, H 4.34, N 9.31.The starting materials are commercially available and were used without further purification. The ligand OPDA  was dissolved in 10 ml of a 1:1 6.Uiso(H) = 1.2Ueq(C) and C—H = 0.93 Å for aromatic H atoms. Hydrogen atoms of the amino groups and of the water molecule were located using a difference-Fourier map and refined with bond-length restraints of 0.89 (1) and 0.85 (1) Å, respectively.Crystal data, data collection and structure refinement details are summarized in Table 210.1107/S2056989023009350/wm5701sup1.cifCrystal structure: contains datablock(s) I. DOI: 10.1107/S2056989023009350/wm5701Isup2.hklStructure factors: contains datablock(s) I. DOI: 2303464CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


Bombina maxima. Here, we reported that B. maxima cells secreted βγ-CAT under glucose, glutamine, and pyruvate deficiency to scavenge extracellular proteins for their nutrient supply and survival. AMPK signaling positively regulated the expression and secretion of βγ-CAT. The PFP complex selectively bound extracellular proteins and promoted proteins uptake through endolysosomal pathways. Elevated intracellular amino acids, enhanced ATP production, and eventually prolonged cell survival were observed in the presence of βγ-CAT and extracellular proteins. Liposome assays indicated that high concentration of ATP negatively regulated the opening of βγ-CAT channels. Collectively, these results uncovered that βγ-CAT is an essential element in cell nutrient scavenging under cell nutrient deficiency by driving vesicular uptake of extracellular proteins, providing a new paradigm for PFPs in cell nutrient acquisition and metabolic flexibility.Nutrient acquisition is essential for animal cells. βγ-CAT is a pore-forming protein (PFP) and trefoil factor complex assembled under tight regulation identified in toad   •The PFP βγ-CAT is expressed and secreted by toad cells under nutrient deficiency•AMPK signaling positively regulates the expression and secretion of βγ-CAT•The secreted βγ-CAT selectively binds and scavenges extracellular nutrients•The high concentrations of ATP may negatively regulate βγ-CAT channel opening Cell biology The former operates in uptake of small nutrient, such as glucose and amino acids, and the latter is the main pathway through which cells obtain insoluble nutrients, such as cholesterol and iron.,,Bombina maxima. It forms membrane pores (channels) with a functional diameter 1.5–2.0 nm.,,,,,Numerous pore-forming proteins (PFPs) with a membrane insertion domain similar to bacterial toxin aerolysin, namely aerolysin family PFPs , have been found in plants and animals.,,,,,This PFP complex firstly targets cell surface acidic glycosphingolipids in lipid rafts via a double-receptor binding model, corresponding to BmALP1 subunit and BmTFF3 subunit binding gangliosides and sulfatides, respectively.B. maxima cells to further investigate the role of SELC protein βγ-CAT in cell nutrient acquisition and metabolic flexibility under cell nutrient deficiency. Interestingly, toad liver and gastrointestinal cells secreted βγ-CAT to scavenge extracellular protein nutrients, such as albumin and ovalbumin, through endolysosomal pathways in the absence of glucose, glutamine and pyruvate, which elevated intracellular amino acids and ATP levels and supported cell survival under nutrient deficiency. The expression and secretion of βγ-CAT were largely attenuated by inhibition of AMP-activated kinase (AMPK) signaling. Furthermore, in a liposome model, ATP, but not AMP inhibited βγ-CAT channel opening. These results revealed the essential role of a secretory PFP in extracellular nutrient scavenging by cells through endolysosomal pathways under cell nutrient deficiency.Here, we used the nutrient deficiency model of toad ,,,,+/Gln+/Pyr+), glucose/glutamine/pyruvate-depleted medium (Glcˉ/Glnˉ/Pyrˉ), and glucose-containing but glutamine/pyruvate-depleted medium (Glc+/Glnˉ/Pyrˉ). In the isolated toad liver cell population, there were 62.4% hepatocytes  on the basis of toad skin transcriptomeT172 and pACC1S80 in human, respectively) were evolutionarily conserved from toad B. maxima to human  in culture supernatants of toad liver cells under Glcˉ/Glnˉ/Pyrˉ conditions , and dextran (70 kDa), an indicator of macropinocytosisKD values of βγ-CAT with OVA and Bm-SA was approximately 2.33 × 10−8 M and 2.47 × 10−8 M, respectively. Ovalbumin-DQ (OVA-DQ) is a fluorescent indicator that fluoresces on proteolytic degradation.ˉ/Glnˉ/Pyrˉ conditions was observed by scanning confocal microscopy  under Glcˉ/Glnˉ/Pyrˉ conditions , toad croscopy B and flocroscopy C. βγ-CATs at 3 h A, a leves βγ-CAT B and 3C.nditions B. Furthenditions D. Ethyl-nditions C and mamnditions D in the nditions E. Collecˉ/Glnˉ/Pyrˉ conditions for 7 h by LC/MS and LC-MS/MS. The levels of amino acids, including asparagine and glutamine, in toad liver cells treated with bovine serum albumin (BSA) and βγ-CAT were augmented compared with those in cells treated with BSA only  staining.nditions A. Notablnditions A. Howevenditions A or ovalnditions B. Additinditions C and 4D.ˉ/Glnˉ/Pyrˉ conditions  on liposomes, which induces dye release from lipid vesicles, an advantageous model without ATP receptors.nditions C. Notablnditions D. This rnditions E. Collec,,B. maxima PFP protein complex βγ-CAT in cell nutrient scavenging and energy supply under the deprivation of glucose and glutamine, essential nutrient components in cell metabolism.B. maxima βγ-CAT indeed acts as a novel system driven cell vesicular delivery that should play a physiological role in cell nutrient acquisition by mediating cellular nutrient import through endolysosomal pathways.,,PFPs are widely distributed in all kingdoms of life, which have long been recognized as either pore-forming toxin for microbial infection or host immune executors.+/Gln+/Pyr+) from 1 to 5 h  and intracellular nutrient sensors, which survey the abundance of energy and major metabolites, play an important role in metabolic homeostasis and cell survival.,,,,in vivo and in vitro in toad osmoregulatory organs to facilitate toad water maintaining.in vivo. Indeed, the secreted βγ-CAT could bind to ovalbumin and toad B. maxima serum albumin (nutrients) directly. Furthermore, the colocalization of βγ-CAT and OVA-DQ was readily observed under cell nutrient deficiency. These results were suggested that the pathway of βγ-CAT importing and enriching nutrients might be selective, which was different from classical pinocytosis/macropinocytosis-like endocytosis.Diverse endocytic pathways are available at the surface of metazoan cells.,,,The functional diameter of bacterial toxin aerolysin channels is approximately 1.5 nm, which is large enough for translocation of oligonucleotides, peptides, and unfolded proteins.,,,,The physiological concentrations of ATP in cells are estimated to be 5–10 mM.,βγ-CAT is a complex of BmALP1 and BmTFF3, in which BmTFF3 acts as a chaperon and regulatory unit of BmALP1 to stabilize the PFP monomer and deliver it to proper targets.,,,It is well documented that autophagy is a cellular process to sequester and degrade intracellular components under nutrient deficiency, which is the last defense for cells under nutrient deficiency and excess autophagy may lead to cell death.B. maxima cells secrete βγ-CAT to scavenge extracellular nutrients under nutrient deficiency at the cellular level (present study). Second, βγ-CAT in toad B. maxima blood circulation is an immediate and active responsive element under toad fasting in vivo, which trans cellularly deliver and transport albumin-bound fatty acids to tissue parenchymal cells for their nutrient supply.B. maxima physiology for adaptation to various nutrient environments. Rationally, similar strategies and executive pathways should be conserved in vertebrates, in which various families of PFPs including af-PFPs are widely distributed. Knowledge from βγ-CAT can provide clues to understand novel PFP-driven cell vesicular delivery systems in nutrient acquisition and metabolic flexibility. Although af-PFPs have not been clearly observed in Eutherian mammals, other PFP family members can readily compensate for the role of af-PFPs.SELC protein βγ-CAT works in cell nutrient acquisition at least at two levels. First, toad B. maxima cells secrete βγ-CAT, a PFP and trefoil factor (TFF) complex assembled depending on environmental cues under glucose, glutamine, and pyruvate deficiency. This PFP complex supports cell survival by driving the cellular import of extracellular proteins through endolysosomal pathways. The imported proteins serve as nutrients in nutrient-deprived cells for energy supply. AMPK signaling positively regulates the expression and secretion of βγ-CAT, whereas high concentrations of ATP (> 1 mM) bind to and negatively regulate βγ-CAT channels. Our findings define the essential role of toad B. maxima PFP complex βγ-CAT in cell macromolecular nutrient scavenging, providing a new paradigm for PFPs in cell nutrient acquisition and metabolic flexibility.In conclusion, the present study elucidated that toad in vitro cell experiments. Consequently, it is necessary to study whether the secretory PFP βγ-CAT contribute to the nutrient acquisition in vivo under starvation, which revealed that the PFP βγ-CAT was secreted into toad blood in response to toad fasting,,,,Our work was restricted zhangy@mail.kiz.ac.cn).Further information and requests for resources and reagents should be directed to and will be fulfilled by the lead contact, Yun Zhang (All unique/stable reagents generated in this study are available from the B. maxima) was performed as described previously and the males with a mean body weight of 25 ± 5 g were used for this study . All procedures and the care and handing of animals were approved by the Ethics Committee of the Kunming Institute of Zoology, Chinese Academy of Sciences .Feeding of toads  containing 10% fetal bovine serum  and 1% Penicillin-Streptomycin Solution  at 37°C with 5% CO+/Gln+/Pyr+ medium, Biological Industries, Cat 01-172-1A), glucose, pyruvate and glutamine-free medium , or added 3.151 g/L glucose to Glcˉ/Glnˉ/Pyrˉ medium (Glc+/Glnˉ/Pyrˉ medium). For macropinocytosis inhibition, cells were first incubated with 100 μM EIPA (5-(N-ethyl-N-isopropyl)-Amiloride), a Na+/H+-exchanger inhibitor used for macropinocytosis inhibition)  in Glcˉ/Glnˉ/Pyrˉ medium for 1 h at 26°C (toad cells) or 37°C (HepG2 cells), respectively. For AMPK signaling inhibition, cells were treated with 0–10 μM compound C or 0–20 μM SBI-0206965  in Glcˉ/Glnˉ/Pyrˉ medium for 3 h at 26°C. To deplete endogenous βγ-CAT, toad cells were incubated with 100 μg/mL anti-βγ-CAT rabbit polyclonal antibodies or 100 μg/mL rabbit IgG  as the isotype control in Glcˉ/Glnˉ/Pyrˉ medium at 26°C.Cells were cultured in basal medium containing 3.151 g/L glucose, 1 mM pyruvate and 2.5 mM glutamine  according to the manufacturer’s instructions.Total 2×106 isolated toad liver cells were cultured at 26°C in various media for 0, 3, 5 and then up to 11 h at 2-h intervals. The treatments were as follows: Glc+/Gln+/Pyr+ medium, Glcˉ/Glnˉ/Pyrˉ medium, Glcˉ/Glnˉ/Pyrˉ medium containing 100 nM βγ-CAT, Glcˉ/Glnˉ/Pyrˉ medium containing 500 μg/mL ovalbumin  or BSA , Glcˉ/Glnˉ/Pyrˉ medium containing 100 nM βγ-CAT and 500 μg/mL OVA or BSA, Glcˉ/Glnˉ/Pyrˉ medium containing 100 μg/mL anti-βγ-CAT antibodies in the presence of 500 μg/mL OVA or BSA, and Glcˉ/Glnˉ/Pyrˉ medium containing 100 μg/mL rabbit IgG in the presence of 500 μg/mL OVA or BSA. To assess the viability of toad stomach cells, 5×106 isolated toad stomach cells were subjected to various treatments  for 7 h at 26°C. To assess cytotoxicity of compound C and SBI-0206965, 5×106 isolated toad liver cells were cultured in Glcˉ/Glnˉ/Pyrˉ medium plus compound C (0–10 μM) or SBI-0206965 (0–20 μM) for 3 h at 26°C. After treatments, toad cells were stained with 500 ng/mL PI  for 4 min at 26°C. Fluorescence was recorded using an LSR Fortessa cell analyzer .Cell viability of toad cells was measured using propidium iodide (PI) stain assay as described previously.6 toad liver cells were cultured in Glc+/Gln+/Pyr+ medium or Glcˉ/Glnˉ/Pyrˉ medium containing various concentration of βγ-CAT (0–1000 nM) for 3 h, then the supernatant of toad cells was collected, and LDH release was detected by LDH cytotoxicity assay kit  according to the manufacturer’s instructions.To assess the cytotoxicity of βγ-CAT in toad liver cells, the total 1×104 cells were seeded into each well of a 96-well culture plate. After overnight culture, the cells were washed thrice with PBS and incubated in Glc+/Gln+/Pyr+ medium, Glcˉ/Glnˉ/Pyrˉ medium, Glcˉ/Glnˉ/Pyrˉ medium plus various concentrations of βγ-CAT (0–160 nM), or Glcˉ/Glnˉ/Pyrˉ medium plus 5 mg/mL OVA in the presence or absence of 40 nM βγ-CAT for 36 h. Then, the cells were incubated with MTS reagent  for 2 h in the dark at 37°C with 5% CO2. Absorbance was read at 490 nm with an Infinite 200 Pro microplate reader .To assess viability of mammalian HepG2 cells, 1×107 toad liver, intestinal, or stomach cells was collected and then washed twice with 30 mL Ringer’s solution. Toad liver cells were cultured in Glc+/Gln+/Pyr+, Glcˉ/Glnˉ/Pyrˉ, or Glc+/Glnˉ/Pyrˉ medium for 1–5 h at 26°C, and intestinal or stomach cells was cultured for 1 h at 26°C. Then, culture supernatants were collected and prepared for the hemolytic activity assay as described previously.A total of 1×10in vitro, and the detail methods were performed as described previouslyKD were determined using Octet Analysis Studio Software using a 1:1 model.The BLI assay was used for the interaction between βγ-CAT and the extracellular nutrients 6 isolated toad liver cells were treated as described in the “B. maxima serum albumin ) or 20 μg/mL Ovalbumin-DQ  in the dark at 26°C for 15 min. Fluorescence was detected by the LSR Fortessa cell analyzer using the FITC channel. In each sample, 1×104 single cells were analyzed.A total of 5×10For immunofluorescence, cells were incubated with 100 μg/mL Ovalbumin-DQ in the dark at 26°C for 15 min. To stain βγ-CAT, cells were washed and incubated with 20 μg/mL anti-βγ-CAT primary antibodies overnight at 4°C in the dark. After washing with PBS three times, the cells were incubated with 10 μg/mL cy3-conjugated anti-rabbit IgG for 1 h at 37°C in the dark. Then, the samples were sealed with an anti-fluorescent quench agent containing DAPI. Images were acquired by a Zeiss LSM 880 microscope system .7 isolated toad liver cells were treated in various media as described in the “2O and MeOH solution (1:4) at −80°C, and then the samples were incubated for 15 min at −80°C. Then, the supernatants were collected by centrifuging at 17,000 g for 10 min 10 μL of these supernatants were injected per analysis. Samples were run on a Vanquish (Thermo Fisher Scientific) UHPLC system with mobile phase A  and mobile phase B (ACN) at a flow rate of 200 μL/min on an Accucore-150-Amide-HILIC column 2.6 μm (100 × 2.1 mm)  at 40°C at a gradient from 45% to 90% A in 15 min followed by a 10-min isocratic step. The UHPLC was coupled to a Q-Exactive (Thermo Fisher Scientific) mass analyzer running in polarity switching mode at 3.5 kV for positive change scanning and 3.0 kV for negative change scanning at an MS1 resolution of 70,000. Metabolites were identified by exact mass (MS1), retention time, and in some cases, by their fragmentation patterns (MS2) at normalized collision energy. Quantification was performed by area under the curve integration of MS1 ion chromatograms with the Thermo Scientific Xcalibur software package. Area values were normalized to cell count averages from triplicate wells treated in parallel for each condition.A total of 1×10Intracellular ATP was extracted and measured by an ATP detection kit  in accordance with the manufacturer’s protocol. Briefly, toad liver cells were treated same as described in the “To assess the effect of adenosine phosphate on βγ-CAT channel formation-induced liposome dye release, liposomes were incubated with 600 nM βγ-CAT in the presence or absence of adenosine phosphate  (Sigma), and then dye release assays were performed as described previously.The purification of βγ-CAT were performed as described previously.KD) was determined using BIA evaluation 4.1 software and its equation for 1:1 Langmuir binding.A direct interaction was assessed as described previously.+/Gln+/Pyr+, Glcˉ/Glnˉ/Pyrˉ, or Glc+/Glnˉ/Pyrˉ medium, respectively. Then, the cells were lysed and prepared for Western blotting. The cell supernatant was concentrated to one-10th of the original volume by a vacuum lyophilizer for Western blotting as described previously.To assess protein level of βγ-CAT in toad liver cell lysate and supernatant, cells were cultured in GlcTo assess AMP-activated protein kinase (AMPK) signaling, cells were treated as described in the “The mRNA levels of BmALP1 (βγ-CAT-α) and BmTFF3 (βγ-CAT-β) in toad liver cells were measured by quantitative real-time PCR (qRT-PCR) using a ChamQ Universal SYBR qPCR Master Mix kit . The cycle counts of target genes were normalized to those of β-actin. Primer sequences of βγ-CAT and β-actin used were the same as report previously.2, and 1 mM EDTA, pH 7.5) for 30 min at 26°C. Cell lysates from HepG2 cells were used as a positive control. Absorbance was read at 620 nm, and the free phosphate concentration was calculated by a standard curve (0–50 μM free phosphate).The ATPase activity of βγ-CAT was measured by an ATPase activity assay kit  according to the manufacturer’s protocol. Briefly, various concentrations βγ-CAT was incubated with 4 mM ATP in assay buffer (40 mM Tris, 80 mM NaCl, 8 mM MgAcB. maxima with other species was conducted by Clustal Omega6.Sequence alignment analysis of AMPKα and ACC1 in t-test. Multiple comparisons were performed by one-way ANOVA with post-hoc contrasts by Dunnett’s multiple comparisons test. Multi-group comparisons were made by two-way ANOVA with Sidak’s multiple comparisons test. p < 0.05 was considered statistically significant. the statistical details of experiments can be found in the figure legends.All experimental values are expressed as means ± SD. Each experiment was repeated at least twice. All data were analyzed using GraphPad Prism 8.0 software. Two comparisons were performed using the standard unpaired


The low-temperature (90 K) crystal structure of 4-(dimethylazaniumyl)-2-hydroxyanilinium dichloride monohydrate is presented along with a Hirshfeld surface analysis of the organic cation. 8H14N2O+·2Cl−·H2O, at low temperature (90 K) are presented. The organic cation is essentially planar: the r.m.s. deviation of its non-hydrogen atoms (aside from the two methyl groups) is 0.0045 Å. The methyl carbons are 1.3125 (12) Å and 1.1278 (12) Å either side of the mean plane. The crystal packing involves extensive hydrogen bonding of types O—H⋯Cl, N—H⋯Cl, N—H⋯OW, and OW—HW⋯Cl (where W = water), which arrange into chains of R24(12) motifs that combine to form corrugated layers parallel to -2-hydroxyanilinium dichloride monohydrate, C Perkin, 1896aka, acetaminophen/Tylenol) and the fenamate family of NSAIDs (anthranilic acid derivatives). Within this context, a concise review of aniline and its derivatives was presented by Anjalin et al. , at 90 K.Given the industrial and pharmaceutical significance of anilinium salts, this paper presents the crystal structure and Hirshfeld-surface analysis of 4-(dimethylazaniumyl)-2-hydroxyanilinium dichloride monohydrate  and N2—H2N⋯Cl2 [D–Ad = 3.0467 (9) Å] hydrogen bonds with the chloride anions, which in turn act as acceptors for O1W—H1W1⋯Cl1vi [D–Ad = 3.1493 (9) Å] and O1W—H2W1⋯Cl2vi [D–Ad = 3.1036 (9) Å] hydrogen bonds with the water molecule (symmetry codes as per Table 1via N1—H3N1⋯O1W [D–Ad = 2.7093 (12) Å] hydrogen bonds, forming chains that extend parallel to [101]  Å] and N1—H1N1⋯Cl2i [D–Ad = 3.1299 (9) Å] hydrogen bonds, forming corrugated layers parallel to  Fig. 3.normd for the cation in I were obtained using CrystalExplorer 4+·4Cl−·4H2O; Stylianou et al., 2017I. Two other anilinium salts not returned in the above search but that share similar features to I are POMXUL  and PAXXIX -(hydroxyimino)methyl]-N,N-dimethyl anilinium chloride (C9H13N2O+·Cl−).A search of the Cambridge Structural Database  and 0.98 Å (RCH3) and Uiso(H) parameters set to either 1.2Ueq (R2CH) or 1.5Ueq (RCH3) of the attached carbon. Nitrogen and oxygen-bound hydrogens were fully refined .Crystal data, data collection and structure refinement details are summarized in Table 210.1107/S2056989023007223/vm2289sup1.cifCrystal structure: contains datablock(s) I, global. DOI: 10.1107/S2056989023007223/vm2289Isup2.hklStructure factors: contains datablock(s) I. DOI: 2289098CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information:


IO2 = 0.209; HIGH FIO2 = 0.155; VHIGH FIO2 = 0.125) on glycaemic control, insulin sensitivity, and oxidative stress during a subsequent oral glucose tolerance test (OGTT) in males with overweight (mean (SD) BMI = 27.6 (1.3) kg/m2; n = 12). Feasibility was defined by exceeding predefined withdrawal criteria for peripheral blood oxygen saturation (SpO2), partial pressure of end‐tidal oxygen or carbon dioxide and acute mountain sickness (AMS), and dyspnoea symptomology. Hypoxia reduced SpO2 in a stepwise manner (CON = 97(1)%; HIGH = 91(1)%; VHIGH = 81(3)%, p < 0.001), but did not affect peak plasma glucose concentration (CON = 7.5(1.8) mmol∙L−1; HIGH = 7.7(1.1) mmol∙L−1; VHIGH = 7.7(1.1) mmol∙L−1; p = 0.777; η2 = 0.013), plasma glucose area under the curve, insulin sensitivity, or metabolic clearance rate of glucose (p > 0.05). We observed no between‐conditions differences in oxidative stress (p > 0.05), but dyspnoea and AMS symptoms increased in VHIGH (p < 0.05), with one participant meeting the withdrawal criteria. Acute HIGH or VHIGH exposure prior to an OGTT does not influence glucose homeostasis in males with overweight, but VHIGH is associated with adverse symptomology and reduced feasibility.Previous research has shown that ≤60 min hypoxic exposure improves subsequent glycaemic control, but the optimal level of hypoxia is unknown and data are lacking from individuals with overweight. We undertook a cross‐over pilot feasibility study investigating the effect of 60‐min prior resting exposure to different inspired oxygen fractions (CON F Likewise, Mackenzie et al.  were reduced, and insulin sensitivity improved, during a subsequent intravenous glucose tolerance test. In men with obesity (mean(SD) body mass index (BMI) = 32.7(1.3) kg/m2), a more prolonged intervention consisting of ten nights of moderate normobaric hypoxia (FIO2 = 0.15) reduced fasting blood [glucose] and improved insulin sensitivity  and two normobaric‐hypoxic conditions equivalent to a simulated altitude of ~2500 m, i.e., ‘high’ altitude , and ~ 4000 m, i.e., ‘very‐high’ altitude   was recruited from the host university (mean (SD) age = 32(7) years; height = 1.82(0.08) m; mass = 91.8(10.1) kg; BMI = 27.6(1.3) kg/m2). Although intended as a pilot feasibility‐study, our sample size was in keeping with similar previous work that informed the study  of 0.14 for a given outcome variable using repeated measures ANOVA . All participants were free from metabolic disease including T2DM (glycated hemoglobin (HbA1c) = 34(4) mmol/mol), thalassemia, and haemoglobinopathies, were not taking any medications and had normal resting lung function , ECG, and blood pressure.To balance the three conditions, a convenience sample of 12 males with overweight  and connected to a metabolic cart  calibrated using known concentrations of O2, CO2, and N2 gas, and a 3‐L air syringe . Participants were blind to the FIO2 on each occasion with CON serving as a sham control condition. Pulmonary gas exchange and ventilation were averaged into 60‐s time bins. Peripheral oxygen saturation was measured by fingertip pulse oximetry  interfaced with a data acquisition system  and heart rate using short range telemetry . Participants were withdrawn if their peripheral blood oxygen saturation (SpO2) fell below 65% for 15 continuous seconds, or if the partial pressure of end‐tidal oxygen (PETO2) fell below 45 mmHg, or the partial pressure of end‐tidal carbon dioxide (PETCO2) fell below 25 mmHg, for three consecutive breaths. Preliminary data collection indicated that, for most participants, the lower FIO2 would approximate the exposure limits permitted within our laboratory.Participants reported to the laboratory at 0730 hours on test days following a 12‐h overnight fast and in a hydrated state, having only consumed water and having abstained from alcohol and strenuous exercise in the previous 24 h. Participants rested in a semi‐recumbent position and a venous cannula was inserted into a forearm vein. Thereafter, following a 15‐min rest period, baseline measurements were obtained and the participant donned an oronasal mask delivering the required FInformed by previous studies showing beneficial effects of acute hypoxia on subsequent glycaemic control  were removed. Energy expenditure and substrate utilization were calculated during the OGTT using indirect respiratory calorimetry, and hunger scores were summed over the OGTT period to create a single hunger score. Criteria for feasibility were (i) withdrawal from the study due to physiological perturbation exceeding any of the study withdrawal criteria for SpO2, PETO2, or PETCO2 or (ii) symptoms attributable to acute hypoxia (AMS and dyspnoea). Between‐conditions differences were examined using one‐way RM‐ANOVA, with the exception of AMS symptoms and perceived dyspnoea, which were analyzed using Friedman's test; if the RM‐ANOVA violated the assumption of sphericity, the Greenhouse–Geisser statistic was employed. The effect size for RM ANOVA is reported as η2, and post‐hoc analysis of significant RM‐ANOVA and Friedman's test effects were undertaken using (least significant difference) pairwise comparisons and Wilcoxon's signed ranks test, respectively.Statistical analyses were undertaken using SPSS Version 28 , and significance was set a priori at P ≤ 0.05. Data are presented mean (SD), with the exception of the Lake Louise questionnaire and dyspnoea scale data, which are presented as median(range). Peak and AUC (trapezoid method) were calculated for [glucose] and AOPP to provide an index of glucose homeostasis and a marker of oxidative stress, respectively, following the hypoxic exposure. Post‐prandial insulin sensitivity and metabolic clearance rate of glucose (a maker of insulin sensitivity) were calculated according to validated equations incorporating demographic data (age and BMI) and on the basis of [glucose] and [insulin] samples at 0, 60, and 120 min of the OGTT  and 0.126(0.05) for the HIGH and VHIGH trials, respectively. SpO2 differed between all conditions (CON = 97(1)%; HIGH = 91(1)%; VHIGH = 81(3)%; p < 0.001, η2 = 0.957) being highest in CON and lowest in VHIGH .One participant was unable to complete the experimental trials due to reaching the study P−1; HIGH = 7.7(1.1) mmol∙L−1; VHIGH = 7.7(1.1) mmol∙L−1; p = 0.777, η2 = 0.013) during the OGTT nor the area under the curve (CON = 11.5(2.9) mmol∙h−1/L; HIGH = 12.3(1.6) mmol∙h−1/L; VHIGH = 12.5(2.0) mmol∙h−1/L; p = 0.241, η2 = 0.146) differed between conditions  μmol∙kg−1∙min−1∙pmol/L; VHIGH = 0.11(0.01) μmol∙kg−1∙min−1∙pmol/L; p = 0.351, η2 = 0.099)  mL∙kg−1∙min−1; VHIGH = 9.6(0.7) mL∙kg−1∙min−1; p = 0.256, η2 = 0.142) were unaffected by the FIO2. Oxidative stress (plasma [AOPP]) before and after the hypoxic intervention period, and during the subsequent OGTT, is shown in Figure p = 0.621, η2 = 0.051) and accumulated oxidative stress (AUC [AOPP]: CON = 526(202) pmol∙h−1/L; HIGH = 532(176) pmol∙h−1/L; VHIGH 567(203) pmol∙h−1/L; p = 0.606, η2 = 0.054) did not differ between the conditions, whereas IL‐6 remained below the detectable limits of the assay (9.4 pg∙mL−1).Plasma [glucose] and [insulin] before and after the hypoxic intervention period, and during the subsequent OGTT, are shown in Figure p = 0.011, η2 = 0.394), being lower in HIGH than VHIGH or CON (p < 0.05), although at an individual substrate level neither the total energy from fat (CON = 104(72) kcal; HIGH = 104(54) kcal; VHIGH = 126(46) kcal; p = 0.220, η2 = 0.155) nor the total energy from carbohydrates (CON = 152(39) kcal; HIGH = 130(46) kcal; VHIGH = 121(30) kcal; p = 0.113, η2 = 0.215) differed between conditions. Similarly, the average RER during the OGTT did not differ between conditions (CON = 0.90(0.08); HIGH = 0.87(0.06); VHIGH = 0.85(0.04); p = 0.092, η2 = 0.233). Hunger score was unaffected by the hypoxic intervention (CON = 220(155) A.U; HIGH = 253(146) A.U; VHIGH = 240(123) A.U; p = 0.331, η2 = 0.116).Total energy expenditure during the OGTT differed between conditions (CON = 256(39) kcal; HIGH = 235(38) kcal; VHIGH = 247(32) kcal; −1; HIGH = 64(6) b∙min−1; VHIGH = 73(10) b∙min−1; p = 0.008, η2 = 0.629 (n = 8)), being higher in both HIGH and VHIGH than CON (both p < 0.01), and higher in VHIGH than HIGH (p < 0.05), but neither systolic blood pressure (CON = 129(10) mmHg; HIGH = 122(9) mmHg; VHIGH = 124(9) mmHg; p = 0.194, η2 = 0.167), diastolic blood pressure (CON = 80(8) mmHg; HIGH = 77(6) mmHg; VHIGH = 78(8) mmHg; p = 0.574, η2 = 0.060), nor mean arterial pressure (CON = 96(8) mmHg; HIGH = 92(7) mmHg; VHIGH = 93(6) mmHg; p = 0.263, η2 = 0.138) differed.Heart rate over the normobaric‐hypoxic exposure period differed between conditions (CON = 61(5) b∙minp = 0.009), which were greater in VHIGH compared to HIGH and CON (p < 0.05), with two participants meeting the criteria for AMS , being higher in VHIGH than HIGH or CON (p < 0.05).There was a between‐conditions difference in AMS symptoms following the normobaric‐hypoxic exposures (CON = 0(0–2); HIGH = 0(0–1); VHIGH = 1(0–7); 4IO2 (VHIGH).The aim of the present study was to assess the short‐term effects of different levels of acute hypoxia on subsequent glucose tolerance, insulin sensitivity, markers of inflammation and oxidative stress, and feasibility in males with overweight. Our main findings were that, following 60 min of breathing either of two hypoxic gas mixtures: (1) neither glucose tolerance, as assessed by peak [glucose] and [glucose] AUC, insulin sensitivity, nor metabolic clearance rate of glucose significantly differed across conditions; (2) markers of oxidative stress and inflammation were not elevated by the acute hypoxic exposures; and (3) feasibility was reduced at the lowest FIO2 of 0.148 did not differ from a normoxic control, and Morishima and Goto (IO2 = 0.155) on resting postprandial glucose regulation. They differ, however, from those showing beneficial effects of acute hypoxia on subsequent glycaemic control in metabolically compromised individuals  indicates that detection of a between‐conditions difference would have been possible with 17 participants .Hypoxia has been shown to increase muscle glucose uptake through effects on insulin‐independent  the highest level of hypoxia that is reliably tolerated without adverse symptoms in individuals with overweight, (ii) the minimal effective exposure duration to this level of hypoxia in this cohort, and (iii) the recruitment of individuals with overweight with overtly compromised glycaemic control.Written informed consent was obtained before participation. The study was approved by the University's Faculty of Science and Health, Research Ethics Committee and conformed to the Declaration of Helsinki, except for registration in a database.


II–dithioether complex. It crystallizes in the monoclinic space group P21/c. Additional Hirshfeld analyses indicate a C—H⋯π interaction along the [010] axis to be the most important packing factor.The title complex represents a further example of a square-planar Pt 2(C7H8I2)2], represents a further example of a square-planar PtII–dithioether complex. It crystallizes in the monoclinic space group P21/c. Additional Hirshfeld analyses indicate a C—H⋯π interaction along the [010] axis to be the most important packing factor.The title complex, [PtI Alternatively, this air-stable complex could be prepared in a much improved yield of 80% by reaction of bis(benzonitrile)diiodoplatinum with 2 equivalents of methyl phenyl sulfide (thioanisol) MeSPh using dichloromethane as solvent. This compound was characterized by NMR spectroscopy in solution and exhibits a singlet resonance for the two magnetically equivalent methyl groups at δ 3.01 ppm, flanked by 195Pt satellites due to a 3JPtH coupling of 48 Hz. Furthermore, we report herein on the solid-state structure and structural analysis of trans-diiodidobis[(methylsulfanyl)benzene-κS]platinum(II) (1). In addition, the results of a Hirshfeld analysis of the intermolecular interactions are presented.In the past, our groups have investigated the coordination of chelating dithioethers such as the vinylic ferrocenyl-dithioether 2.trans-Diiodidobis[(methylsulfanyl)benzene-κS]platinum(II) (1) crystallizes from dichloromethane in the monoclinic crystal system, space group P21/c. The molecular structure of 1 is presented in Figs. 21, which shows Ch2 symmetry. The distance from the coordinating iodine center I1 to Pt1 is 2.61205 (15) Å, showing a slight elongation with respect to its educt structure trans-[PtI2(NCPh)2] (2) (2.6052 (8) Å; Viola et al., 2018trans-[PtCl2(SMePh)2] (3) reported by Ahlgrén 2] (4) 2] (5)  Å in 5]. The chelate complexes cis-diiodo-platinum(II) diiodoplatinum(II) °, C2—S1—Pt1 = 104.52 (7)° and C2—S1—C1 = 103.46 (11)°.All further bonds have characteristic dimensions  x, y, −z] with a distance between the phenyl ring and H1Bi of 2.5377 (10) Å benzene] ligand and its oxidized derivative are focused on now. The already compared structure trans-dichloro-bis[methyl(phenyl)sulfanyl]platinum palladium has been published independently by two different research groups [JISWUD platinum(II) platinum(II) 2SiPh2}] 2SiPh2}]·DCM 2Si2Me4}] 2Si2Me4}]·DCM benzene]platinum (1) was synthesized by adding methylphenyl sulfide  dissolved in 0.5 mL of dichloromethane via a microsyringe to a solution of bis(benzonitrile)diiodoplatinum  in dichloromethane (3 mL) and stirring overnight at room temperature. trans-Diiodobis[(methylsulfanyl)benzene]platinum  was isolated as red crystals after layering with heptane.14H16I2PtS2 (697.30 g mol−1): C, 24.11; H, 2.32; S, 9.20. Found: C, 23.92; H, 2.21; S, 9.05%.Calculated for C1H NMR : δ = 3.01 , 7.05–7.73  ppm.6.Uiso(H) = 1.2Ueq(C) for CH2 and CH hydrogen atoms and Uiso (H) = 1.5Ueq(C) for CH3 hydrogen atoms.Crystal data, data collection and structure refinement details are summarized in Table 210.1107/S2056989023003717/jy2030sup1.cifCrystal structure: contains datablock(s) I, global. DOI: 10.1107/S2056989023003717/jy2030Isup2.hklStructure factors: contains datablock(s) I. DOI: 2258409CCDC reference:  crystallographic information; 3D view; checkCIF reportAdditional supporting information: