Predicted deleterious variants in ABCA1, LPL, LPA and KIF6 are associated with statin response and adverse events in patients with familial hypercholesterolemia and disturb protein structure and stability.

OBJECTIVES: This study explored the association of deleterious variants in pharmacodynamics (PD) genes with statin response and adverse effects in patients with familial hypercholesterolemia (FH) and analyzed their potential effects on protein structure and stability. METHODS: Clinical and laboratory data were obtained from 144 adult FH patients treated with statins. A panel of 32 PD genes was analyzed by exon-targeted gene sequencing. Deleterious variants were identified using prediction algorithms and their structural effects were analyzed by molecular modeling studies. RESULTS: A total of 102 variants were predicted as deleterious (83 missense, 8 stop-gain, 4 frameshift, 1 indel, 6 splicing). The variants ABCA1 rs769705621 (indel), LPA rs41267807 (p.Tyr2023Cys) and KIF6 rs20455 (p.Trp719Arg) were associated with reduced low-density lipoprotein cholesterol (LDLc) response to statins, and the LPL rs1801177 (p.Asp36Asn) with increased LDLc response (P < 0.05). LPA rs3124784 (p.Arg2016Cys) was predicted to increase statin response (P = 0.022), and ABCA1 rs769705621 to increase the risk of statin-related adverse events (SRAE) (P = 0.027). LPA p.Arg2016Cys and LPL p.Asn36Asp maintained interactions with solvent, LPA p.Tyr2023Cys reduced intramolecular interaction with Gln1987, and KIF6 p.Trp719Arg did not affect intramolecular interactions. DDMut analysis showed that LPA p.Arg2016Cys and p.Tyr2023Cys and LPL p.Asp36Asn caused energetically favorable changes, and KIF6 p.Trp719Arg resulted in unfavorable energetic changes, affecting protein stability. CONCLUSION: Deleterious variants in ABCA1, LPA, LPL and KIF6 are associated with variability in LDLc response to statins, and ABCA1 rs769705621 is associated with SRAE risk in FH patients. Molecular modeling studies suggest that LPA p.Tyr2023Cys and KIF6 p.Trp719Arg disturb protein conformational structure and stability.

Exploring the dark side of tertiary and quaternary structure dynamics in MtbFBPaseII.

Tuberculosis (TB) is a major global cause of mortality, primarily stemming from latent tuberculosis infection (LTBI). Failure to fully treat LTBI can result in drug-resistant forms of TB. Therefore, it is essential to develop novel drugs with unique mechanisms of action to combat TB effectively. One crucial metabolic pathway in Mycobacterium tuberculosis (Mtb), which contributes to TB infection and persistence, is gluconeogenesis. Within this pathway, the enzyme fructose bisphosphatase (FBPase) plays a significant role and is considered a promising target for drug development. By targeting MtbFBPaseII, a specific class of FBPase, researchers have employed molecular dynamics simulations to identify regions capable of binding new drugs, thereby inhibiting the enzyme's activity and potentially paving the way for the development of effective treatments.Communicated by Ramaswamy H. Sarma.

Innovative Microemulsion Loaded with Unusual Dimeric Flavonoids from Fridericia platyphylla (Cham.) L.G. Lohmann Roots.

Fridericia platyphylla (Cham.) L.G. Lohmann is a species native to the Brazilian cerrado, with promising bioactivity. The organic fraction of the roots is rich in unusual dimeric flavonoids, reported as potential candidates for cancer treatment. The exploration of these flavonoids is very important, considering their diverse biological activities and the need for innovative therapeutic options. This work aimed to develop and characterize a microemulsion loaded with a non-polar fraction (DCM). The constituents were chosen, and the pseudo-ternary diagram was constructed to determine the region of microemulsion formation. The microemulsions blank (ME), with 3% (ME3) and 5% (ME5) of fraction DCM, were characterized in terms of droplet size, zeta potential, and polydispersity index. Both MEs showed particle sizes <100 nm; only ME3 exhibited better values for polydispersity index and zeta potential and was therefore selected for further study. The organoleptic and physicochemical characteristics were evaluated, revealing limpidity and transparency typical of these microstructures, physiologically acceptable pH, refractive index of 1.42±0.01, and density of 1.017 g/cm3±0.01. The stability tests showed good stability profiles even after exposure to extreme thermal conditions, with minimal changes in pH and the content of the incorporated fraction. The in vitro release study demonstrated that ME3 enabled the controlled release of the fraction, with a cumulative amount released over 60% within 6 h. Furthermore, fraction DCM and ME3 exhibited no toxicity in Tenebrio molitor larvae. The developed microemulsion exhibited excellent properties, so this study represents the first successful attempt to develop a formulation that incorporates the dimeric flavonoid fraction.

Covalent Inhibitors for Neglected Diseases: An Exploration of Novel Therapeutic Options.

Neglected diseases, primarily found in tropical regions of the world, present a significant challenge for impoverished populations. Currently, there are 20 diseases considered neglected, which greatly impact the health of affected populations and result in difficult-to-control social and economic consequences. Unfortunately, for the majority of these diseases, there are few or no drugs available for patient treatment, and the few drugs that do exist often lack adequate safety and efficacy. As a result, there is a pressing need to discover and design new drugs to address these neglected diseases. This requires the identification of different targets and interactions to be studied. In recent years, there has been a growing focus on studying enzyme covalent inhibitors as a potential treatment for neglected diseases. In this review, we will explore examples of how these inhibitors have been used to target Human African Trypanosomiasis, Chagas disease, and Malaria, highlighting some of the most promising results so far. Ultimately, this review aims to inspire medicinal chemists to pursue the development of new drug candidates for these neglected diseases, and to encourage greater investment in research in this area.

Exploring the conformational dynamics of the CRD4: a model for receptor lysosomal activity shifts in search of 'sweet' and 'sour' states.

The macrophage mannose receptor (RMM) is a crucial component of the immune system involved in immune responses, inflammation resolution, and tissue remodeling. When RMM is activated by a specific ligand, it undergoes internalization, forming an endosome that matures into a lysosome. Within the lysosome, structural changes in RMM facilitate the dissociation of ligands for further processing. However, the precise details of these structural changes are not well understood. In this study, we used molecular dynamics simulations to investigate the conformational dynamics of a specific region called CRD4 in RMM. Our simulations explored different conditions, including pH variations and the presence of Ca2+ ions. By analyzing the simulation data, we found that conformational changes primarily occur in loop regions, while the secondary structure remains stable. The binding site of CRD4, essential for ligand interaction, is located on the protein surface between two specific loop regions. Ligand binding is stabilized by three important amino acids. Interestingly, the interaction patterns differ between monosaccharide and disaccharide ligands. These findings improve our understanding of CRD4's dynamics and how it recognizes ligands. They provide insights into the structure of CRD4 and its role in ligand dissociation within lysosomes. The study also highlights the significance of loop regions in functional dynamics and interactions. Further research is needed to fully uncover the complete structure of CRD4, understand ligand binding modes, and explore the influence of environmental factors. This study lays the foundation for future investigations targeting carbohydrate-protein interactions and the development of therapeutics based on RMM's unique properties.Communicated by Ramaswamy H. Sarma.

Identification of pathogenic variants in the Brazilian cohort with Familial hypercholesterolemia using exon-targeted gene sequencing.

Familial hypercholesterolemia (FH) is a monogenic disease characterized by high plasma low-density lipoprotein cholesterol (LDL-c) levels and increased risk of premature atherosclerotic cardiovascular disease. Mutations in FH-related genes account for 40% of FH cases worldwide. In this study, we aimed to assess the pathogenic variants in FH-related genes in the Brazilian FH cohort FHBGEP using exon-targeted gene sequencing (ETGS) strategy. FH patients (n = 210) were enrolled at five clinical sites and peripheral blood samples were obtained for laboratory testing and genomic DNA extraction. ETGS was performed using MiSeq platform (Illumina). To identify deleterious variants in LDLR, APOB, PCSK9, and LDLRAP1, the long-reads were subjected to Burrows-Wheeler Aligner (BWA) for alignment and mapping, followed by variant calling using Genome Analysis Toolkit (GATK) and ANNOVAR for variant annotation. The variants were further filtered using in-house custom scripts and classified according to the American College Medical Genetics and Genomics (ACMG) guidelines. A total of 174 variants were identified including 85 missense, 3 stop-gain, 9 splice-site, 6 InDel, and 71 in regulatory regions (3'UTR and 5'UTR). Fifty-two patients (24.7%) had 30 known pathogenic or likely pathogenic variants in FH-related genes according to the American College Medical and Genetics and Genomics guidelines. Fifty-three known variants were classified as benign, or likely benign and 87 known variants have shown uncertain significance. Four novel variants were discovered and classified as such due to their absence in existing databases. In conclusion, ETGS and in silico prediction studies are useful tools for screening deleterious variants and identification of novel variants in FH-related genes, they also contribute to the molecular diagnosis in the FHBGEP cohort.

Dynamics of the personalities of PCSK9 on missense variants (rs505151 and rs562556) from elderly cohort studies in Brazil.

The Proprotein Convertase Subtilisin/Kexin Type 9 (PCSK9) promotes the degradation of the low-density lipoprotein receptors (LDLR). Gain-of-function (GOF) variants of PCSK9 significantly affects lipid metabolism leading to coronary artery disease (CAD), owing to the raising the plasma low-density lipoprotein (LDL). Considering the public health matter, large-scale genomic studies have been conducted worldwide to provide the genetic architecture of populations for the implementation of precision medicine actions. Nevertheless, despite the advances in genomic studies, non-European populations are still underrepresented in public genomic data banks. Despite this, we found two high-frequency variants (rs505151 and rs562556) in the ABraOM databank (Brazilian genomic variants) from a cohort SABE study conducted in the largest city of Brazil, São Paulo. Here, we assessed the structural and dynamical features of these variants against WT through a molecular dynamics study. We sought fundamental dynamical interdomain relations through Perturb Response Scanning (PRS) and we found an interesting change of dynamical relation between prodomain and Cysteine-Histidine-Rich-Domain (CHRD) in the variants. The results highlight the pivotal role of prodomain in the PCSK9 dynamic and the implications for the development of new drugs depending on patient group genotype.

Trypanosoma cruzi Sirtuin 2 as a Relevant Druggable Target: New Inhibitors Developed by Computer-Aided Drug Design.

Trypanosoma cruzi, the etiological agent of Chagas disease, relies on finely coordinated epigenetic regulation during the transition between hosts. Herein we targeted the silent information regulator 2 (Sir2) enzyme, a NAD+-dependent class III histone deacetylase, to interfere with the parasites' cell cycle. A combination of molecular modelling with on-target experimental validation was used to discover new inhibitors from commercially available compound libraries. We selected six inhibitors from the virtual screening, which were validated on the recombinant Sir2 enzyme. The most potent inhibitor (CDMS-01, IC50 = 40 µM) was chosen as a potential lead compound.

LDLR missense variants disturb structural conformation and LDLR activity in T-lymphocytes of Familial hypercholesterolemia patients.

Familial hypercholesterolemia (FH) is caused by deleterious mutations in the LDLR that increase markedly low-density lipoprotein (LDL) cholesterol and cause premature atherosclerotic cardiovascular disease. Functional effects of pathogenic LDLR variants identified in Brazilian FH patients were assessed using in vitro and in silico studies. Variants in LDLR and other FH-related genes were detected by exon-target gene sequencing. T-lymphocytes were isolated from 26 FH patients, and 3 healthy controls and LDLR expression and activity were assessed by flow cytometry and confocal microscopy. The impact of LDLR missense variants on protein structure was assessed by molecular modeling analysis. Ten pathogenic or likely pathogenic LDLR variants (six missense, two stop-gain, one frameshift, and one in splicing region) and six non-pathogenic variants were identified. Carriers of pathogenic and non-pathogenic variants had lower LDL binding and uptake in activated T-lymphocytes compared to controls (p < 0.05), but these variants did not influence LDLR expression on cell surface. Reduced LDL binding and uptake was also observed in carriers of LDLR null and defective variants. Modeling analysis showed that p.(Ala431Thr), p.(Gly549Asp) and p.(Gly592Glu) disturb intramolecular interactions of LDLR, and p.(Gly373Asp) and p.(Ile488Thr) reduce the stability of the LDLR protein. Docking and molecular interactions analyses showed that p.(Cys184Tyr) and p.(Gly373Asp) alter interaction of LDLR with Apolipoprotein B (ApoB). In conclusion, LDLR null and defective variants reduce LDL binding capacity and uptake in activated T-lymphocytes of FH patients and LDLR missense variants affect LDLR conformational stability and dissociation of the LDLR-ApoB complex, having a potential role in FH pathogenesis.

Effects of PCSK9 missense variants on molecular conformation and biological activity in transfected HEK293FT cells.

PCSK9 gain-of-function (GOF) variants increase degradation of low-density lipoprotein receptor (LDLR) and are potentially associated with Familial Hypercholesterolemia (FH). This study aimed to explore the effects of PCSK9 missense variants on protein structure and interactions with LDLR using molecular modeling analyses and in vitro functional studies. Variants in FH-related genes were identified in a Brazilian FH cohort using an exon-target gene sequencing strategy. Eight PCSK9 missense variants in pro- [p.(E32K) and p.(E57K)], catalytic [p.(R237W), p.(P279T) and p.(A443T)], and C-terminal histidine-cysteine rich (CHR) [p.(R469W), p.(Q619P) and p.(R680Q)] domains were identified. Molecular dynamics analyses revealed that GOF variants p.(E32K) and p.(R469W) increased extreme motions in PCSK9 amino acid backbone fluctuations and affected Hbond and water bridge interactions between the pro-domain and CM1 region of the CHR domain. HEK293FT cells transfected with plasmids carrying p.(E32K) and p.(R469W) variants reduced LDLR expression (8.7 % and 14.8 %, respectively) compared to wild type (p < 0.05) but these GOF variants did not affect PCSK9 expression and secretion. The missense variants p.(P279T) and p.(Q619P) also reduced protein stability and altered Hbond interactions. In conclusion, PCSK9 p.(E32K), p.(R469W), p.(P279T) and p.(Q619P) variants disrupt intramolecular interactions that are essential for PCSK9 structural conformation and biological activity and may have a potential role in FH pathogenesis.

Do Go Chasing Waterfalls: Enoyl Reductase (FabI) in Complex with Inhibitors Stabilizes the Tetrameric Structure and Opens Water Channels.

The enzyme enoyl-ACP reductase (FabI) is the limiting step of the membrane's fatty acid biosynthesis in bacteria and a druggable target for novel antibacterial agents. The FabI active form is a homotetramer, which displays the highest affinity to inhibitors. Herein, molecular dynamics studies were carried out using the structure of FabI in complex with known inhibitors to investigate their effects on tetramerization. Our results suggest that multimerization is essential for the integrity of the catalytic site and that inhibitor binding enables the multimerization by stabilizing the substrate binding loop (SBL, L:195-200) coupled with changes in the H4/5 (QR interface). We also observed that AFN-1252 (naphtpyridinone derivative) promotes unique conformational changes affecting monomer-monomer interfaces. These changes are induced by AFN-1252 interaction with key residues in the binding sites (Ala95, Tyr146, and Tyr156). In addition, the analysis of water trajectories indicated that AFN-1252 complexes allow more water molecules to enter the binding site than triclosan and MUT056399 complexes. FabI-AFN-1252 simulations show accumulation of water molecules near the Tyr146/147 pocket, which can become a hotspot to the design of novel FabI inhibitors.

Zika Virus NS2B-NS3 Protease: Quantum Chemical Properties Insights into Designing Inhibitory Peptides.

BACKGROUND: Zika fever affects poor and vulnerable populations, presenting cycles observed in, at least 86 countries, with no vaccine prevention or treatment available. It is known that the genus Flavivirus causes Zika Virus (ZIKV), as Dengue and Yellow Fever, whose genetic material decodes, among other proteins, a series of non-structural (NS) proteins essential for viral replication, such as NS2B-NS3 protease. Additionally, chemical and biological systems are commonly studied using molecular modeling approaches allowing, among several other processes, to elucidate mechanisms of action, molecule reactivity and/or chemical properties and the design of new drugs. Thus, considering the in silico complexes between the biological target and the bioactive molecule, it is possible to understand better experimental results based on molecular properties, which are compared with the findings of the biological activity. OBJECTIVE: Accordingly, this study aimed to present computational docking simulations of five previously reported active peptides against NS2B-NS3 protease of ZIKV and analyze some quantum chemical properties to identify the main contribution to improving the action. METHODS: The compounds were described by Rut and coworkers (2017) and Hill and coworkers (2018), submitted to docking simulation in Gold software and quantum chemical properties calculations in Wavefunction Spartan software. RESULTS: Total energy, electrophilicity index (ω) and energy gap (GAP) appeared to be the best properties to justify the peptide's biological activity. Moreover, the most promising compound (P1, Km 4.18 µM) had the best value of total energy (- 2763.04001 au), electrophilicity index (8.04 eV) and GAP (6.49 eV), indicating an energetically favorable molecule with good interaction with the target and, when compared to other peptides, presented moderate reactivity. P4 showed the highest electrophilicity index value (28.64 eV), which justified the interaction ability visualized in the docking simulation. However, its GAP value (4.24 eV) was the lowest in the series, suggesting high instability, possibly validating its low biological activity value (Km 19 uM). GAP was important to understand the chemical instability, and high values can promote damage to biological response. CONCLUSION: Furthermore, it was also noted that high electron affinity, related to the electrophilicity index, promoted electron-accepting characteristics, which was important to improve the biological activity of the peptides. A larger compound series must be studied to access features more precisely. However, these results have paramount importance in guiding future effort in this extremely-need health area.

Inhibitor induced conformational changes in SARS-COV-2 papain-like protease.

SARS-CoV-2's papain-like protease (PLpro) interaction with ligands has recently been explored with a myriad of crystal structures. We used molecular dynamics (MD) simulations to study different PLpro-ligand complexes, their ligand-induced conformational changes, and interactions. We focused on inhibitors reported with known IC50 against PLpro, namely GRL-0617, XR8-89, PLP_Snyder530, and Sander's recently published compound 7 (CPD7), and compared these trajectories against the apostructure (Apo), with a total of around 60 µs worth simulation data. We aimed to study the conformational changes using molecular dynamics simulations for the inhibitors in the PLpro. PCA analyses and the MSM models revealed distinct conformations of PLpro in the absence/presence of ligands and proposed that BL2-loop contributes to the accessibility of these inhibitors. Further, bulkier substituents closer to Tyr268 and Gln269 could improve inhibition of SARS-CoV-2 PLpro by occupying the region between BL2-groove and BL2-loop, but we also expand on the relevance of exploring multiple PLpro sub-pockets to improve inhibition.

Genetic Variant ABCC1 rs45511401 Is Associated with Increased Response to Statins in Patients with Familial Hypercholesterolemia.

Statins are the first-line treatment for familial hypercholesterolemia (FH), but response is highly variable due to genetic and nongenetic factors. Here, we explored the association between response and genetic variability in 114 Brazilian adult FH patients. Specifically, a panel of 84 genes was analyzed by exon-targeted gene sequencing (ETGS), and the functional impact of variants in pharmacokinetic (PK) genes was assessed using an array of functionality prediction methods. Low-density lipoprotein cholesterol (LDL-c) response to statins (reduction ≥ 50%) and statin-related adverse event (SRAE) risk were assessed in carriers of deleterious variants in PK-related genes using multivariate linear regression analyses. Fifty-eight (50.8%) FH patients responded to statins, and 24 (21.0%) had SRAE. Results of the multivariate regression analysis revealed that ABCC1 rs45511401 significantly increased LDL-c reduction after statin treatment (p < 0.05). In silico analysis of the amino-acid change using molecular docking showed that ABCC1 rs45511401 possibly impairs statin efflux. Deleterious variants in PK genes were not associated with an increased risk of SRAE. In conclusion, the deleterious variant ABCC1 rs45511401 enhanced LDL-c response in Brazilian FH patients. As such, this variant might be a promising candidate for the individualization of statin therapy.

Development, Optimization, and Validation of Forensic Analytical Method for Quantification of Anticholinesterase Pesticides in Biological Matrices from Suspected Cases of Animal Poisoning.

Anticholinesterase pesticides are a main cause of the intentional or accidental poisoning of animals. Anticholinesterases include several substances that cause the overstimulation of both central and peripheral acetylcholine-dependent neurotransmission. Forensic analyses of poisoning cases require high levels of expertise, are costly, and often do not provide reliable quantitative information for unambiguous conclusions. The purpose of the present study was to develop and validate a method of high-performance liquid chromatography with diode array detector (HPLC−DAD) for the identification and quantitation of n-methyl carbamates, organophosphates and respective metabolites from biological samples of animals that were suspected of poisoning. HPLC−DAD is reliable, fast, simplistic and cost-effective. The method was validated for biological samples obtained from stomach contents, liver, vitreous humor and blood from four different animal species. The validation of the method was achieved using the following analytical parameters: linearity, precision, accuracy, selectivity, recovery, and matrix effect. The method showed linearity at the range of 25−500 µg/mL, and the correlation coefficient (r2) values were >0.99 for all matrices. Precision and accuracy were determined by the (a) coefficient of variation (CV), (b) relative standard deviation low-quality control (LQC), (c) medium-quality control (QCM), and (d) high-quality control (QCA). The indicated parameters were all less than 15%. The recovery of analytes ranged from 31 to 71%. The analysis of results showed no significant interfering peaks due to common xenobiotics or matrix effects. The abovementioned method was used to positively identify pesticide analytes in 44 of the 51 animal samples that were suspected of poisoning, demonstrating its usefulness as a forensic tool.

Rational Design, Synthesis, and Mechanism of (3S,4R)-3-Amino-4-(difluoromethyl)cyclopent-1-ene-1-carboxylic Acid: Employing a Second-Deprotonation Strategy for Selectivity of Human Ornithine Aminotransferase over GABA Aminotransferase.

Human ornithine aminotransferase (hOAT) is a pyridoxal 5'-phosphate (PLP)-dependent enzyme that contains a similar active site to that of γ-aminobutyric acid aminotransferase (GABA-AT). Recently, pharmacological inhibition of hOAT was recognized as a potential therapeutic approach for hepatocellular carcinoma. In this work, we first studied the inactivation mechanisms of hOAT by two well-known GABA-AT inactivators (CPP-115 and OV329). Inspired by the inactivation mechanistic difference between these two aminotransferases, a series of analogues were designed and synthesized, leading to the discovery of analogue 10b as a highly selective and potent hOAT inhibitor. Intact protein mass spectrometry, protein crystallography, and dialysis experiments indicated that 10b was converted to an irreversible tight-binding adduct (34) in the active site of hOAT, as was the unsaturated analogue (11). The comparison of kinetic studies between 10b and 11 suggested that the active intermediate (17b) was only generated in hOAT and not in GABA-AT. Molecular docking studies and pKa computational calculations highlighted the importance of chirality and the endocyclic double bond for inhibitory activity. The turnover mechanism of 10b was supported by mass spectrometric analysis of dissociable products and fluoride ion release experiments. Notably, the stopped-flow experiments were highly consistent with the proposed mechanism, suggesting a relatively slow hydrolysis rate for hOAT. The novel second-deprotonation mechanism of 10b contributes to its high potency and significantly enhanced selectivity for hOAT inhibition.

Carnitine palmitoyl transferase I: Conformational changes induced by long-chain fatty acyl CoA ligands.

The Carnitine Palmitoyltranferase I (CPT1) catalyzes the rate-limiting step of long-chain fatty acid (LCFA) mitochondrial ß-oxidation. The enzyme promotes the conjugation of LCFA with l-carnitine, which allows LCFA to enter the mitochondria matrix. The structural features involved in CPT1 and LCFA-CoA interactions have not been fully elucidated, mainly due to the absence of CPT1 crystallographic data. Previous studies reported important residues (Lys556, Lys560, and Lys561) crucial to the CPT1 mechanism. Nonetheless, these studies have not explored the LCFA bindings. Using molecular modeling strategies, we aimed to understand the conformational changes in CPT1 structure induced by LCFA-CoA. For this purpose, a tridimensional CPT1A model was built by homology modeling using CRAT protein (PBD:1t7q, resolution 1.8 Å) as a template. We simulated the CPT1 structure in the presence and absence of LCFA-CoA by molecular dynamics (MD). By applying a principal component analysis (PCA), two states of apostructure CPT1 based on CoA-Loop (688-711) were observed. In contrast, just one state was evidenced along with smaller conformational subspaces in ligand-complexed simulations using LCFA-CoA. The CoA moiety of ligands interacts with charged residues, namely Lys560, Lys556, Arg563, and Arg645. The frequency of interactions observed for each of these residues is <60% of simulation time, suggesting a dynamic profile of interactions in synergy with long-chain carbon interactions over α-I (478-492). Collectively, these features may be associated with the catalytic conformation of LCFA-CoA to CPT1a. Further calculations of free-energy for different fatty acids, such as alpha-linolenic (ALA), gamma-linolenic (GLA), and arachidonic (ARA) acids, yielded energy values ranging from -76.9 ± 15.9 to -68.5 ± 10.0 kcal mol-1. In conclusion, the present structural model and simulations provide molecular-level insights into LCFA-CoA and CPT1a interactions. These findings may help to further knowledge on the conformational changes of CPT1a induced by LCFA-CoA derivates.

SARS-COV-2 Mpro conformational changes induced by covalently bound ligands.

SARS-CoV-2's main protease (Mpro) interaction with ligands has been explored with a myriad of crystal structures, most of the monomers. Nonetheless, Mpro is known to be active as a dimer but the relevance of the dimerization in the ligand-induced conformational changes has not been fully elucidated. We systematically simulated different Mpro-ligand complexes aiming to study their conformational changes and interactions, through molecular dynamics (MD). We focused on covalently bound ligands (N1 and N3, ∼9 µs per system both monomers and dimers) and compared these trajectories against the apostructure. Our results suggest that the monomeric simulations led to an unrealistically flexible active site. In contrast, the Mpro dimer displayed a stable oxyanion-loop conformation along the trajectory. Also, ligand interactions with residues His41, Gly143, His163, Glu166 and Gln189 are postulated to impact the ligands' inhibitory activity significantly. In dimeric simulations, especially Gly143 and His163 have increased interaction frequencies. In conclusion, long-timescale MD is a more suitable tool for exploring in silico the activity of bioactive compounds that potentially inhibit the dimeric form of SARS-CoV-2 Mpro.Communicated by Ramaswamy H. Sarma.