Photocatalytic Conversion of CO2 to Formate/CO by an (η6-para-Cymene)Ru(II) Half-Metallocene Catalyst: Influence of Additives and TiO2 Immobilization on the Catalytic Mechanism and Product Selectivity.

The catalytic efficacy of the monobipyridyl (η6-para-Cymene)Ru(II) half-metallocene, [(p-Cym)Ru(bpy)Cl]+ was evaluated in both mixed homogeneous (dye + catalyst) and heterogeneous hybrid systems (dye/TiO2/Catalyst) for photochemical CO2 reduction. A series of homogeneous photolysis experiments revealed that the (p-Cym)Ru(II) catalyst engages in two competitive routes for CO2 reduction (CO2 to formate conversion via RuII-hydride vs CO2 to CO conversion through a RuII-COOH intermediate). The conversion activity and product selectivity were notably impacted by the pKa value and the concentration of the proton source added. When a more acidic TEOA additive was introduced, the half-metallocene Ru(II) catalyst leaned toward producing formate through the RuII-H mechanism, with a formate selectivity of 86%. On the other hand, in homogeneous catalysis with TFE additive, the CO2-to-formate conversion through RuII-H was less effective, yielding a more efficient CO2-to-CO conversion with a selectivity of >80% (TONformate of 140 and TONCO of 626 over 48 h). The preference between the two pathways was elucidated through an electrochemical mechanistic study, monitoring the fate of the metal-hydride intermediate. Compared to the homogeneous system, the TiO2-heterogenized (p-Cym)Ru(II) catalyst demonstrated enhanced and enduring performance, attaining TONs of 1000 for CO2-to-CO and 665 for CO2-to-formate.

Perfluorooctanoic acid inhibits cell proliferation through mitochondrial damage.

Grown evidence has shown that the liver and reproductive organs were the main target organs of perfluorooctanoic acid (PFOA). Herein, we studied a toxic mechanism of PFOA using HeLa Chang liver epithelial cells. When incubated with PFOA for 24 h or 48 h, cell proliferation was inhibited in a concentration- and time-dependent fashion, but interestingly, the feature of dead cells was not notable. Mitochondrial volume was increased with concentration and time, whereas the mitochondrial membrane potential and produced ATP amounts were significantly reduced. Autophagosome-like vacuoles and contraction of the mitochondrial inner membrane were observed in PFOA-treated cells. The expression of acetyl CoA carboxylase (ACC) and p-ACC proteins rapidly decreased, and that of mitochondrial dynamics-related proteins increased. The expression of solute carrier family 7 genes, ChaC glutathione-specific gamma-glutamylcyclotransferase 1, and 5S ribosomal RNA gene was up-regulated the most in cells exposed to PFOA for 24 h, and the KEGG pathway analysis revealed that PFOA the most affected metabolic pathways and olfactory transduction. More importantly, PPAR alpha, fatty acid binding protein 1, and CYP450 family 1 subfamily A member 1 were identified as the target proteins for binding between PFOA and cells. Taken together, we suggest that disruption of mitochondrial integrity and function may contribute closely to PFOA-induced cell proliferation inhibition.

Elucidating TH301's influence on the torsion angle of CRY1 W399 using replica exchange with solute tempering (REST) molecular dynamics (MD) simulations.

Cryptochrome 1 (CRY1) is a protein involved in the circadian clock and associated with various diseases. Targeting CRY1 for drug development requires the discovery of competitive inhibitors that target its FAD binding site through ubiquitination. During the development of compounds to regulate CRY1, an intriguing compound called TH301 was identified. Despite binding to CRY1, TH301 does not induce the expected reaction and is considered an inactive compound. However, it has been observed that TH301 affects the torsion angle of CRY1's W399 residue, which plays a crucial role in the regulation of ubiquitination by influencing the movement of the lid loop. In our research, we aimed to understand how TH301 induces the torsion angle of CRY1's W399 to shift to an "out-form" by performing REST-based MD simulations. The cyclopentane of TH301 tends to align parallel with W292, creating a repulsive force when W399 is in the "in-form", leading to a flip. In the "out-form", W399's side chain interacts with TH301's chlorobenzene through a π-π interaction, stabilizing this pose. This analysis helps identify compounds binding to CRY1 and filter out inactive ones. We found that assessing the interaction energy between TH301 and W399 is crucial to evaluate whether W399 flips or not. These findings contribute to the development of drugs targeting CRY1 and enhance our understanding of its regulatory mechanisms.

Comparison of disinfectants-induced gene expression profile: Potential adverse effects.

Despite their importance in combating the spread of the COVID-19 pandemic, adverse effects of disinfectants on human health, especially the respiratory system, have been of continuing concern to researchers. Considering that bronchi are the main target of sprayed disinfectants, we here treated the seven major active ingredients in disinfectant products accepted by the US EPA to human bronchial epithelial cells and determined the subtoxic levels. Then, we performed microarray analysis using total RNA obtained at the subtoxic level and designed a network representing disinfectant-induced cellular response using the KEGG pathway analysis technique. Polyhexamethylguanidine phosphate, a lung fibrosis inducer, was used as a reference material to verify the relationship between cell death and pathology. The derived results reveal potential adverse effects along with the need for an effective application strategy for each chemical.

Combining QM/MM Calculations with Classical Mining Minima to Predict Protein-Ligand Binding Free Energy.

We developed an effective binding free energy prediction protocol which incorporates quantum mechanical/molecular mechanical (QM/MM) calculations to substitute the specified atomic charges of force fields with quantum-mechanically recalculated ones at a proposed pose using a mining minima approach with the VeraChem mining minima engine. We tested this protocol using seven well-known targets with 147 different ligands and compared it with classical mining minima and the most popular binding free energy (BFE) methods using different metrics. Our new protocol, dubbed Qcharge-VM2, yielded an overall Pearson correlation of 0.86, which was better than all the methods examined. Qcharge-VM2 performed significantly better than implicit solvent-based methods, such as MM-GBSA and MM-PBSA, but not as good as explicit water-based free energy perturbation methods, such as FEP+, in terms of root-mean-square error, RMSE (1.75 kcal/mol) and mean unsigned error, MUE (1.39 kcal/mol) on a limited set of targets. However, our protocol is substantially less computationally demanding compared with FEP+. The combined accuracy and efficiency of our method can be valuable in drug discovery campaigns.

Sensitization of GSH synthesis by curcumin curtails acrolein-induced alveolar epithelial apoptosis via Keap1 cysteine conjugation: A randomized controlled trial and experimental animal model of pneumonitis.

INTRODUCTION: Epidemiological studies have reported an association between exposures to ambient air pollution and respiratory diseases, including chronic obstructive pulmonary disease (COPD). Pneumonitis is a critical driving factor of COPD and exposure to air pollutants (e.g., acrolein) is associated with increased incidence of pneumonitis. OBJECTIVES: Currently available anti-inflammatory therapies provide little benefit against respiratory diseases. To this end, we investigated the preventive role of curcumin against air pollutant-associated pneumonitis and its underlying mechanism. METHODS: A total of 40 subjects was recruited from Chengdu, China which is among the top three cities in terms of respiratory mortality related to air pollution. The participants were randomly provided either placebo or curcumin supplements for 2 weeks and blood samples were collected at the baseline and at the end of the intervention to monitor systemic markers. In our follow up mechanistic study, C57BL/6 mice (n = 40) were randomly allocated into 4 groups: Control group (saline + no acrolein), Curcumin only group (curcumin + no acrolein), Acrolein only group (saline + acrolein), and Acrolein + Curcumin group (curcumin + acrolein). Curcumin was orally administered at 100 mg/kg body weight once a day for 10 days, and then the mice were subjected to nasal instillation of acrolein (5 mg/kg body weight). Twelve hours after single acrolein exposure, all mice were euthanized. RESULTS: Curcumin supplementation, with no noticeable adverse responses, reduced circulating pro-inflammatory cytokines in association with clinical pneumonitis as positive predictive while improving those of anti-inflammatory cytokines. In the pre-clinical study, curcumin reduced pneumonitis manifestations by suppression of intrinsic and extrinsic apoptotic signaling, which is attributed to enhanced redox sensing of Nrf2 and thus sensitized synthesis and restoration of GSH, at least in part, through curcumin-Keap1 conjugation. CONCLUSIONS: Our study collectively suggests that curcumin could provide an effective preventive measure against air pollutant-enhanced pneumonitis and thus COPD.

High-temperature adaptation of an OsNRT2.3 allele is thermoregulated by small RNAs.

Climate change negatively affects crop yield, which hinders efforts to reach agricultural sustainability and food security. Here, we show that a previously unidentified allele of the nitrate transporter gene OsNRT2.3 is required to maintain high yield and high nitrogen use efficiency under high temperatures. We demonstrate that this tolerance to high temperatures in rice accessions harboring the HTNE-2 (high temperature resistant and nitrogen efficient-2) alleles from enhanced translation of the OsNRT2.3b mRNA isoform and the decreased abundance of a unique small RNA (sNRT2.3-1) derived from the 5' untranslated region of OsNRT2.3. sNRT2.3-1 binds to the OsNRT2.3a mRNA in a temperature-dependent manner. Our findings reveal that allelic variation in the 5' untranslated region of OsNRT2.3 leads to an increase in OsNRT2.3b protein levels and higher yield during high-temperature stress. Our results also provide a breeding strategy to produce rice varieties with higher grain yield and lower N fertilizer input suitable for a sustainable agriculture that is resilient against climate change.

Identification of highly selective type II kinase inhibitors with chiral peptidomimetic tails.

Identification of highly selective type II kinase inhibitors is described. Two different chiral peptidomimetic scaffolds were introduced on the tail region of non-selective type II kinase inhibitor GNF-7 to enhance the selectivity. Kinome-wide selectivity profiling analysis showed that type II kinase inhibitor 7a potently inhibited Lck kinase with great selectivity (IC50 of 23.0 nM). It was found that 7a and its derivatives possessed high selectivity for Lck over even structurally conserved all Src family kinases. We also observed that 7a inhibited Lck activation in Jurkat T cells. Moreover, 7a was found to alleviate clinical symptoms in DSS-induced colitis mice. This study provides a novel insight into the design of selective type II kinase inhibitors by adopting chiral peptidomimetic moieties on the tail region.

Effect of Structural Fine-Tuning on Chelate Stability and Liver Uptake of Anionic MRI Contrast Agents.

The purpose of this study is to assess the physicochemical properties and MRI diagnostic efficacy of two newly synthesized 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA)-type Gd chelates, Gd-SucL and Gd-GluL, with an asymmetric α-substituted pendant arm as potential hepatocyte-specific magnetic resonance imaging contrast agents (MRI CAs). Our findings show that fine conformational changes in the chelating arm affect the in vivo pharmacokinetic behavior of the MRI CA, and that a six-membered chelating substituent of Gd-SucL is more advantageous in this system to avoid unwanted interactions with endogenous species. Gd-SucL exhibited a general DOTA-like chelate stability trend, indicating that all chelating arms retain coordination bonding. Finally, the in vivo diagnostic efficacy of highly stable Gd-SucL as a potential hepatocyte-specific MRI CA was evaluated using T1-weighted MR imaging on an orthotopic hepatocarcinoma model.

Rare Missense Functional Variants at COL4A1 and COL4A2 in Sporadic Intracerebral Hemorrhage.

ObjectiveTo test the genetic contribution of rare missense variants in COL4A1 and COL4A2 in which common variants are genetically associated with sporadic intracerebral hemorrhage (ICH), we performed rare variant analysis in multiple sequencing data for the risk for sporadic ICH.MethodsWe performed sequencing across 559Kbp at 13q34 including COL4A1 and COL4A2 among 2,133 individuals (1,055 ICH cases; 1,078 controls) in US-based and 1,492 individuals (192 ICH cases; 1,189 controls) from Scotland-based cohorts, followed by sequence annotation, functional impact prediction, genetic association testing, and in silico thermodynamic modeling.ResultsWe identified 107 rare nonsynonymous variants in sporadic ICH, of which two missense variants, rs138269346 (COL4A1I110T) and rs201716258 (COL4A2H203L), were predicted to be highly functional and occurred in multiple ICH cases but not in controls from the US-based cohort. The minor allele of rs201716258 was also present in Scottish ICH patients, and rs138269346 was observed in two ICH-free controls with a history of hypertension and myocardial infarction. Rs138269346 was nominally associated with non-lobar ICH risk (P=0.05), but not with lobar ICH (P=0.08), while associations between rs201716258 and ICH subtypes were non-significant (P>0.12). Both variants were considered pathogenic based on minor allele frequency (<0.00035 in EUR), predicted functional impact (deleterious or probably damaging), and in silico modeling studies (substantially altered physical length and thermal stability of collagen).ConclusionsWe identified rare missense variants in COL4A1/A2 in association with sporadic ICH. Our annotation and simulation studies suggest that these variants are highly functional and may represent targets for translational follow-up.

pH-promoted O-α-glucosylation of flavonoids using an engineered α-glucosidase mutant.

Retaining glycosidase mutants lacking its general acid/base catalytic residue are originally termed thioglycoligases which synthesize thio-linked disaccharides using sugar acceptor bearing a nucleophilic thiol group. A few thioglycoligases derived from retaining α-glycosidases have been classified into a new class of catalysts, O-glycoligases which transfer sugar moiety to a hydroxy group of sugar acceptors, resulting in the formation of O-linked glycosides or oligosaccharides. In this study, an efficient O-α-glucosylation of flavonoids was developed using an O-α-glycoligase derived from a thermostable α-glucosidase from Sulfolobus solfataricus (MalA-D416A). The O-glycoligase exhibited efficient transglycosylation activity with a broad substrate spectrum for all kinds of tested flavonoids including flavone, flavonol, flavanone, flavanonol, flavanol and isoflavone classes in yields of higher than 90%. The glucosylation by MalA-D416A preferred alkaline conditions, suggesting that pH-promoted deprotonation of hydroxyl groups of the flavonoids would accelerate turnover of covalent enzyme intermediate via transglucosylation. More importantly, the glucosylation of flavonoids by MalA-D416A was exclusively regioselective, resulting in the synthesis of flavonoid 7-O-α-glucosides as the sole product. Kinetic analysis and molecular dynamics simulations provided insights into the acceptor specificity and the regiospecificity of O-α-glucosylation by MalA-D416A. This pH promoted transglycosylation using O-α-glycoligases may prove to be a general synthesis route to flavonoid O-α-glycosides.

Target-Specific Drug Design Method Combining Deep Learning and Water Pharmacophore.

Following identification of a target protein, hit identification, which finds small organic molecules that bind to the target, is an important first step of a structure-based drug design project. In this study, we demonstrate a target-specific drug design method that can autonomously generate a series of target-favorable compounds. This method utilizes the seq2seq model based on a deep learning algorithm and a water pharmacophore. Water pharmacophore models are used to screen compounds that are favorable to a given target in a large compound database, and seq2seq compound generators are used to train the screened compounds and generate entirely new compounds based on the training model. Our method was tested through binding energy calculation studies of six pharmaceutically relevant targets in the directory of useful decoys (DUD) set with docking. The compounds generated by our method had lower average binding energies than decoy compounds in five out of six cases and included a number of compounds that had lower binding energies than the average binding energies of the active compounds in four cases. The generated compound lists for these four cases featured compounds with lower binding energies than even the most active compounds.

Elucidation of Mechanism for Ligand Efficacy at Leukotriene B4 Receptor 2 (BLT2).

G protein-coupled receptors (GPCRs) have always been important drug targets in the pharmaceutical industry. One major question for the current GPCR drug discovery is how drugs have distinct efficacies at the same GPCR target. Related to this question, we studied how different ligands can have disparate efficacies at Leukotriene B4 receptor (BLT2). By using molecular modeling studies, we predicted that Tyr2716.51 located at TM6 of BLT2 performs as a key trigger for its activation and verified the prediction by site-directed mutagenesis, chemotactic motility studies, which included a chemical derivative of agonist CAY10583. We further identified Asn2756.55 located at TM6 as a weak activation trigger in BLT2 and performed double mutation studies to confirm our computational results. Our results provide strong evidence for the exact mechanism of ligand efficacy at BLT2.

Gadolinium-Based Neuroprognostic Magnetic Resonance Imaging Agents Suppress COX-2 for Prevention of Reperfusion Injury after Stroke.

Advancements in recanalization therapies have rendered reperfusion injury an important challenge for stroke management. It is essential to work toward effective therapeutics that protect the ischemic brain from reperfusion injury. Here, we report a new concept of neuroprognostic agents, which combine molecular diagnostic imaging and targeted neuroprotection for treatment of reperfusion injury after stroke. These neuroprognostic agents are inflammation-targeted gadolinium compounds conjugated with nonsteroidal anti-inflammatory drugs (NSAIDs). Our results demonstrated that gadolinium-based MRI contrast agents conjugated with NSAIDs suppressed the increase in cyclooxygenase-2 (COX-2) levels, ameliorated glial activation, and neuron damage that are phenotypic for stroke by mitigating neuroinflammation, which prevented reperfusion injury. In addition, this study showed that the neuroprognostic agents are promising T1 molecular MRI contrast agents for detecting precise reperfusion injury locations at the molecular level. Our results build on this new concept of neuroprognostics as a novel management strategy for ischemia-reperfusion injury, combining neuroprotection and molecular diagnostics.

Influence of alkyl chain substitution of ammonium ionic liquids on the activity and stability of tobacco etch virus protease.

Ionic liquids (ILs) are known to provide stability to biomolecules. ILs are also widely used in the fields of chemical engineering, biological engineering, chemistry, and biochemistry because they facilitate enzyme catalyzed reactions and enhance their conversion rate. In this work, we have evaluated the influence of alkyl chain substitution of ammonium ILs such as diethylammonium dihydrogen phosphate (DEAP) and triethylammonium hydrogen phosphate (TEAP) for the stability and activity of the tobacco etch virus (TEV) protease. Further, we performed molecular dynamics (MD) simulations to calculate the RMSD (root mean square deviation) for TEV and TEV + ILs. Experimental and simulations results show that TEV is more stable in the presence of TEAP than DEAP. Whereas, TEV protease activity for the cleavage of fusion proteins is preserved in the presence of DEAP while lost in the presence of TEAP. Hence, DEAP IL can serve as alternative solvents for the stability of the TEV protease with preserved activity. To the best of our knowledge, this is first study to show that ILs can stabilize and maintain the TEV protease cleavage activity.

Elucidation of the inhibition mechanism of sulfiredoxin using molecular modeling and development of its inhibitors.

When intracellular reactive oxygen species (ROS) increase, cancer cells are more vulnerable to oxidative stress compared to normal cells; thus, the collapse of redox homeostasis can lead to selective death of cancer cells. Indeed, recent studies have shown that inhibition of sulfiredoxin (Srx), which participates in antioxidant mechanisms, induces ROS-mediated cancer cell death. In this paper, we describe how an Srx inhibitor, J14 (4-[[[4-[4-(2-chlor-ophenyl)-1-piperazinyl]-6-phenyl-2-pyrimidinyl]thio]methyl]-benzoic acid), interferes with the antioxidant activity of Srx at the molecular level. We searched for possible binding sites of Srx using a binding site prediction method and uncovered two possible inhibition mechanisms of Srx by J14. Using molecular dynamics simulations and binding free energy calculations, we confirmed that J14 binds to the ATP binding site; therefore, J14 acts as a competitive inhibitor of ATP, settling the question of the two mechanisms. Based on the inhibition mechanism revealed at the atomic level, we designed several derivatives of J14, which led to LMT-328 (4-(((4-(4-(2-Chlorophenyl)piperazin-1-yl)-6-(2,4-dihydroxy-5-isopropylphenyl)pyrimidin-2-yl)thio)methyl)benzoic acid), which is possibly an even more potent inhibitor than J14.

Englerin A-sensing charged residues for transient receptor potential canonical 5 channel activation.

The transient receptor potential canonical (TRPC) 5 channel, known as a nonselective cation channel, has a crucial role in calcium influx. TRPC5 has been reported to be activated by muscarinic receptor activation and extracellular pH change and inhibited by the protein kinase C pathway. Recent studies have also suggested that TRPC5 is extracellularly activated by englerin A (EA), but the mechanism remains unclear. The purpose of this study is to identify the EA-interaction sites in TRPC5 and thereby clarify the mechanism of TRPC5 activation. TRPC5 channels are over-expressed in human embryonic kidney (HEK293) cells. TRPC5 mutants were generated by site-directed mutagenesis. The whole-cell patch-clamp configuration was used to record TRPC5 currents. Western analysis was also performed to observe the expression of TRPC5 mutants. To identify the EA-interaction site in TRPC5, we first generated pore mutants. When screening the mutants with EA, we observed the EA-induced current increases of TRPC5 abolished in K554N, H594N, and E598Q mutants. The current increases of other mutants were reduced in different levels. We also examined the functional intactness of the mutants that had no effect by EA with TRPC5 agonists, such as carbachol or GTPγS. Our results suggest that the three residues, Lys-554, His-594, and Glu-598, in TRPC5 might be responsible for direct interaction with EA, inducing the channel activation. We also suggest that although other pore residues are not critical, they could partly contribute to the EA-induced channel activation.

Ginsenoside Rd Inhibits the Metastasis of Colorectal Cancer via Epidermal Growth Factor Receptor Signaling Axis.

Ginsenoside Rd is a saponin from ginseng and has been reported to have various biological activities. However, the effect of ginsenoside Rd on the metastasis of colorectal cancer (CRC) remains unknown. Here, we found that ginsenoside Rd decreased the colony-forming ability, migration, invasion, and wound-healing abilities of CRC cells, although it did not affect cell proliferation. In addition, using an inverse-docking assay, we found that ginsenoside Rd bound to epidermal growth factor receptor (EGFR) with a high binding affinity, inducing the downregulation of stemness- and epithelial-mesenchymal transition-related genes; these were partially rescued by either exogenous EGF treatment or ectopic expression of SOX2. Furthermore, ginsenoside Rd significantly decreased the number and size of tumor metastasis nodules in the livers, lungs, and kidneys of mouse model of metastasis. © 2018 IUBMB Life, 71(5):601-610, 2019.

Synthesis and Evaluation of Manganese(II)-Based Ethylenediaminetetraacetic Acid-Ethoxybenzyl Conjugate as a Highly Stable Hepatobiliary Magnetic Resonance Imaging Contrast Agent.

In this study, we designed and synthesized a highly stable manganese (Mn2+)-based hepatobiliary complex by tethering an ethoxybenzyl (EOB) moiety with an ethylenediaminetetraacetic acid (EDTA) coordination cage as an alternative to the well-established hepatobiliary gadolinium (Gd3+) chelates and evaluated its usage as a T1 hepatobiliary magnetic resonance imaging (MRI) contrast agent (CA). This new complex exhibits higher r1 relaxivity (2.3 mM-1 s-1) than clinically approved Mn2+-based hepatobiliary complex Mn-DPDP (1.6 mM-1 s-1) at 1.5 T. Mn-EDTA-EOB shows much higher kinetic inertness than that of clinically approved Gd3+-based hepatobiliary MRI CAs, such as Gd-DTPA-EOB and Gd-BOPTA. In addition, in vivo biodistribution and MRI enhancement patterns of this new Mn2+ chelate are comparable to those of Gd3+-based hepatobiliary MRI CAs. The diagnostic efficacy of the new complex was demonstrated by its enhanced tumor detection sensitivity in a liver cancer model using in vivo MRI.