Molecular dynamics of mismatch detection-How MutS uses indirect readout to find errors in DNA.

The mismatch repair protein MutS safeguards genomic integrity by finding and initiating repair of basepairing errors in DNA. Single-molecule studies show MutS diffusing on DNA, presumably scanning for mispaired/unpaired bases, and crystal structures show a characteristic "mismatch-recognition" complex with DNA enclosed within MutS and kinked at the site of error. But how MutS goes from scanning thousands of Watson-Crick basepairs to recognizing rare mismatches remains unanswered, largely because atomic-resolution data on the search process are lacking. Here, 10 µs all-atom molecular dynamics simulations of Thermus aquaticus MutS bound to homoduplex DNA and T-bulge DNA illuminate the structural dynamics underlying the search mechanism. MutS-DNA interactions constitute a multistep mechanism to check DNA over two helical turns for its 1) shape, through contacts with the sugar-phosphate backbone, 2) conformational flexibility, through bending/unbending engineered by large-scale motions of the clamp domain, and 3) local deformability, through basepair destabilizing contacts. Thus, MutS can localize a potential target by indirect readout due to lower energetic costs of bending mismatched DNA and identify a site that distorts easily due to weaker base stacking and pairing as a mismatch. The MutS signature Phe-X-Glu motif can then lock in the mismatch-recognition complex to initiate repair.

A computational strategy for therapeutic development against superoxide dismutase (SOD1) amyloid formation: effect of polyphenols on the various events in the aggregation pathway.

Pathology of superoxide dismutase 1 (SOD1) aggregation is linked to a neurodegenerative disease known as amyotrophic lateral sclerosis (ALS). Without suitable post-translational modifications (PTMs), the protein structure tends to become aggregation-prone. Understanding the role of PTMs and targeting the aggregation-prone SOD1 with small molecules can be used to design a strategy to inhibit its aggregation. Microsecond long molecular dynamics (MD) simulations followed by free energy surface (FES) analyses show that the loss of structure in the apo monomer happens locally and stepwise. Removing the disulfide bond from apoprotein leads to further instability in the zinc-binding loop, giving rise to non-native protein conformations. Further, it was found that these non-native conformations have a higher propensity to form a non-native dimer. We chose three structurally similar polyphenols based on their binding energies and investigated their impact on SOD1 aggregation kinetics. MD simulations of apo-SOD1SH/corkscrew fibril-polyphenol complexes were also carried out. The effect of polyphenols was seen on fibril elongation as well. Based on the experiments and MD simulation results, it can be inferred that the choice of inhibitors is influenced not only by the binding energy but also by dimer interface stabilization, the proclivity to form non-native dimers, the propensity to break fibrils, and the propensity to decrease the rate of elongation. The polyphenols with 3' and 4' hydroxyl groups are better inhibitors of SOD1 aggregation.

Stimulation of heat shock protein 90 chaperone function through binding of a novobiocin analog KU-32.

Heat shock protein 90 (Hsp90) is a eukaryotic chaperone responsible for the folding and functional activation of numerous client proteins, many of which are oncoproteins. Thus, Hsp90 inhibition has been intensely pursued, resulting in the development of many potential Hsp90 inhibitors, not all of which are well-characterized. Hsp90 inhibitors not only abrogate its chaperone functions, but also could help us gain insight into the structure-function relationship of this chaperone. Here, using biochemical and cell-based assays along with isothermal titration calorimetry, we investigate KU-32, a derivative of the Hsp90 inhibitor novobiocin (NB), for its ability to modulate Hsp90 chaperone function. Although NB and KU-32 differ only slightly in structure, we found that upon binding, they induce completely opposite conformational changes in Hsp90. We observed that NB and KU-32 both bind to the C-terminal domain of Hsp90, but surprisingly, KU-32 stimulated the chaperone functions of Hsp90 via allosteric modulation of its N-terminal domain, responsible for the chaperone's ATPase activity. In vitro and in silico studies indicated that upon KU-32 binding, Hsp90 undergoes global structural changes leading to the formation of a "partially closed" intermediate that selectively binds ATP and increases ATPase activity. We also report that KU-32 promotes HeLa cell survival and enhances the refolding of an Hsp90 substrate inside the cell. This discovery explains the effectiveness of KU-32 analogs in the management of neuropathies and may facilitate the design of molecules that promote cell survival by enhancing Hsp90 chaperone function and reducing the load of misfolded proteins in cells.

Structural Insights into the DNA-Binding Mechanism of BCL11A: The Integral Role of ZnF6.

The transcription factor BCL11A is a critical regulator of the switch from fetal hemoglobin (HbF: α 2 γ 2 ) to adult hemoglobin (HbA: α 2 ß 2 ) during development. BCL11A binds at a cognate recognition site (TGACCA) in the γ-globin gene promoter and represses its expression. DNA-binding is mediated by a triple zinc finger domain, designated ZnF456. Here, we report comprehensive investigation of ZnF456, leveraging X-ray crystallography and NMR to determine the structures in both the presence and absence of DNA. We delve into the dynamics and mode of interaction with DNA. Moreover, we discovered that the last zinc finger of BCL11A (ZnF6) plays a special role in DNA binding and γ-globin gene repression. Our findings help account for some rare γ-globin gene promoter mutations that perturb BCL11A binding and lead to increased HbF in adults (hereditary persistence of fetal hemoglobin). Comprehending the DNA binding mechanism of BCL11A opens avenues for the strategic, structure-based design of novel therapeutics targeting sickle cell disease and ß-thalassemia.

Structural insights into the DNA-binding mechanism of BCL11A: The integral role of ZnF6.

The transcription factor BCL11A is a critical regulator of the switch from fetal hemoglobin (HbF: α2γ2) to adult hemoglobin (HbA: α2ß2) during development. BCL11A binds at a cognate recognition site (TGACCA) in the γ-globin gene promoter and represses its expression. DNA-binding is mediated by a triple zinc finger domain, designated ZnF456. Here, we report comprehensive investigation of ZnF456, leveraging X-ray crystallography and NMR to determine the structures in both the presence and absence of DNA. We delve into the dynamics and mode of interaction with DNA. Moreover, we discovered that the last zinc finger of BCL11A (ZnF6) plays a different role compared to ZnF4 and 5, providing a positive entropic contribution to DNA binding and γ-globin gene repression. Comprehending the DNA binding mechanism of BCL11A opens avenues for the strategic, structure-based design of novel therapeutics targeting sickle cell disease and ß-thalassemia.

Ergothioneine boosts mitochondrial respiration and exercise performance via direct activation of MPST.

Ergothioneine (EGT) is a diet-derived, atypical amino acid that accumulates to high levels in human tissues. Reduced EGT levels have been linked to age-related disorders, including neurodegenerative and cardiovascular diseases, while EGT supplementation is protective in a broad range of disease and aging models in mice. Despite these promising data, the direct and physiologically relevant molecular target of EGT has remained elusive. Here we use a systematic approach to identify how mitochondria remodel their metabolome in response to exercise training. From this data, we find that EGT accumulates in muscle mitochondria upon exercise training. Proteome-wide thermal stability studies identify 3-mercaptopyruvate sulfurtransferase (MPST) as a direct molecular target of EGT; EGT binds to and activates MPST, thereby boosting mitochondrial respiration and exercise training performance in mice. Together, these data identify the first physiologically relevant EGT target and establish the EGT-MPST axis as a molecular mechanism for regulating mitochondrial function and exercise performance.

In silico study of interaction of (ZnO)12 nanocluster to glucose oxidase-FAD in absence and presence of glucose.

Diabetes mellitus is one of the foremost global concerns, as it has impacted millions of lives. Therefore, there is an urgent need to develop a technology for continuous glucose monitoring in vivo. In the current study, we employed computational methods such as docking, MD simulations, and MM/GBSA, to obtain molecular insights into the interaction between (ZnO)12 nanocluster and glucose oxidase (GOx) that cannot be obtained through experiments alone. For this, theoretical modeling of the 3D cage-like (ZnO)12 nanocluster in ground state configuration was performed. Further docking of (ZnO)12 nanocluster with GOx molecule was carried out to find the nano-bio-interaction of (ZnO)12-GOx complex. To understand the whole interaction and dynamics of (ZnO)12-GOx-FAD-with and without glucose, we performed MD simulation and MM/GBSA analysis of (ZnO)12-GOx-FAD complex and glucose-(ZnO)12-GOx-FAD complex separately. The interaction was found to be stable, and the binding energy of (ZnO)12 to GOx-FAD increases in the presence of glucose by 6 kcal mol-1. This may be helpful in nano probing of the interaction of GOx with glucose. It can help in making a device like fluorescence resonance energy transfer (FRET) based nano-biosensor to monitor the glucose level in pre and post diabetic patient.Communicated by Ramaswamy H. Sarma.

Approaches and Perspective of Coarse-Grained Modeling and Simulation for Polymer-Nanoparticle Hybrid Systems.

Molecular modeling and simulations have emerged as effective and indispensable tools to characterize polymeric systems. They provide fundamental and essential insights to design a product of the required properties and to improve the understanding of a phenomenon at the molecular level for a particular system. The polymer-nanoparticle hybrids are materials with outstanding properties and correspondingly large applications whose study has benefited from this new paradigm. However, despite the significant expansion of modern day computational powers, investigation of the long time and large length scale phenomenon in polymeric and polymer-nanoparticle systems is still a challenging task to complete through all-atom molecular dynamics (AA-MD) simulations. To circumvent this problem, a variety of coarse-grained (CG) models have been proposed, ranging from the generic CG models for qualitative properties predictions to more realistic chemically specific CG models for quantitative properties predictions. These CG models have already delivered some success stories in the study of several spatial and temporal evolutions of many processes. Some of these studies were beyond the feasibility of traditional atomistic resolution models due to either the size or the time constraints. This review captures the different types of popular CG approaches that are utilized in the investigation of the microscopic behavior of polymer-nanoparticle hybrid systems. The rationale of this article is to furnish an overview of the popular CG approaches and their applications, to review several important and most recent developments, and to delineate the perspectives on future directions in the field.

Emerging frontiers in virtual drug discovery: From quantum mechanical methods to deep learning approaches.

Virtual screening-based approaches to discover initial hit and lead compounds have the potential to reduce both the cost and time of early drug discovery stages, as well as to find inhibitors for even challenging target sites such as protein-protein interfaces. Here in this review, we provide an overview of the progress that has been made in virtual screening methodology and technology on multiple fronts in recent years. The advent of ultra-large virtual screens, in which hundreds of millions to billions of compounds are screened, has proven to be a powerful approach to discover highly potent hit compounds. However, these developments are just the tip of the iceberg, with new technologies and methods emerging to propel the field forward. Examples include novel machine-learning approaches, which can reduce the computational costs of virtual screening dramatically, while progress in quantum-mechanical approaches can increase the accuracy of predictions of various small molecule properties.

Designed thiazolidines: an arsenal for the inhibition of nsP3 of CHIKV using molecular docking and MD simulations.

Chikungunya virus (CHIKV) belongs to the alpha virus and it's infection in humans causes fever, known as chikungunya fever (CHIKF). It is a sudden onset of fever and may affect humans badly. The mode of transmission to human occurs due to the biting of the mosquitoes. Till date, thousands of humans are affected from this virus throughout the world. As on date, no promising medicine or vaccine is available in the market to cure from this viral infection. Therefore, there is a need of promising candidate against the nsp3 of CHIKV. In the present work, a library of the compounds are designed based on the product obtained in a multi-component reaction. Then, the designed compounds are filtered based on binding energy against the nsp3 of CHIKV obtained using molecular docking. Further, to understand the interaction of nsp3 of CHIKV and screened compound, CMPD474, the molecular dynamics (MD) simulations at different temperatures, that is, 300, 325, 350, 375, and 400 K is performed. The binding or the formation of the complex is studied through different trajectories obtained from MD simulations. The accurate information for the binding energy is determined by performing MM-GBSA calculations and the best inhibition was observed at 300 K as the change in free energy for the formation of the complex is -7.0523 kcal/mol.Communicated by Ramaswamy H. Sarma.

Understanding the binding affinity of noscapines with protease of SARS-CoV-2 for COVID-19 using MD simulations at different temperatures.

The current outbreak of a novel coronavirus, named as SARS-CoV-2 causing COVID-19 occurred in 2019, is in dire need of finding potential therapeutic agents. Recently, ongoing viral epidemic due to coronavirus (SARS-CoV-2) primarily affected mainland China that now threatened to spread to populations in most countries of the world. In spite of this, there is currently no antiviral drug/ vaccine available against coronavirus infection, COVID-19. In the present study, computer-aided drug design-based screening to find out promising inhibitors against the coronavirus (SARS-CoV-2) leads to infection, COVID-19. The lead therapeutic molecule was investigated through docking and molecular dynamics simulations. In this, binding affinity of noscapines(23B)-protease of SARS-CoV-2 complex was evaluated through MD simulations at different temperatures. Our research group has established that noscapine is a chemotherapeutic agent for the treatment of drug resistant cancers; however, noscapine was also being used as anti-malarial, anti-stroke and cough-suppressant. This study suggests for the first time that noscapine exerts its antiviral effects by inhibiting viral protein synthesis.

Estradiol induces allosteric coupling and partitioning of sex-hormone-binding globulin monomers among conformational states.

Sex-hormone-binding globulin (SHBG) regulates the transport and bioavailability of estradiol. The dynamics of estradiol's binding to SHBG are incompletely understood, although it is believed that estradiol binds to each monomer of SHBG dimer with identical affinity (Kd ∼2 nM). Contrary to the prevalent view, we show that estradiol's binding to SHBG is nonlinear, and the "apparent" Kd changes with varying estradiol and SHBG concentrations. Estradiol's binding to each SHBG monomer influences residues in the ligand-binding pocket of both monomers and differentially alters the conformational and energy landscapes of both monomers. Monomers are not energetically or conformationally equivalent even in fully bound state. Estradiol's binding to SHBG involves bidirectional, inter-monomeric allostery that changes the distribution of both monomers among various energy and conformational states. Inter-monomeric allostery offers a mechanism to extend the binding range of SHBG and regulate hormone bioavailability as estradiol concentrations vary widely during life.

Promising inhibitors of main protease of novel corona virus to prevent the spread of COVID-19 using docking and molecular dynamics simulation.

Coronavirus disease-2019 (COVID-19) is a global health emergency and the matter of serious concern, which has been declared a pandemic by WHO. Till date, no potential medicine/ drug is available to cure the infected persons from SARS-CoV-2. This deadly virus is named as novel 2019-nCoV coronavirus and caused coronavirus disease, that is, COVID-19. The first case of SARS-CoV-2 infection in human was confirmed in the Wuhan city of the China. COVID-19 is an infectious disease and spread from man to man as well as surface to man . In the present work, in silico approach was followed to find potential molecule to control this infection. Authors have screened more than one million molecules available in the ZINC database and taken the best two compounds based on binding energy score. These lead molecules were further studied through docking against the main protease of SARS-CoV-2. Then, molecular dynamics simulations of the main protease with and without screened compounds were performed at room temperature to determine the thermodynamic parameters to understand the inhibition. Further, molecular dynamics simulations at different temperatures were performed to understand the efficiency of the inhibition of the main protease in the presence of the screened compounds. Change in energy for the formation of the complexes between the main protease of novel coronavirus and ZINC20601870 as well ZINC00793735 at room temperature was determined on applying MM-GBSA calculations. Docking and molecular dynamics simulations showed their antiviral potential and may inhibit viral replication experimentally. Communicated by Ramaswamy H. Sarma.

Allosterically Coupled Multisite Binding of Testosterone to Human Serum Albumin.

Human serum albumin (HSA) acts as a carrier for testosterone, other sex hormones, fatty acids, and drugs. However, the dynamics of testosterone's binding to HSA and the structure of its binding sites remain incompletely understood. Here, we characterize the dynamics of testosterone's binding to HSA and the stoichiometry and structural location of the binding sites using 2-dimensional nuclear magnetic resonance (2D NMR), fluorescence spectroscopy, 4,4'-dianilino-1,1'-binaphthyl-5,5'-disulfonic acid dipotassium salt partitioning, and equilibrium dialysis, complemented by molecular modeling. 2D NMR studies showed that testosterone competitively displaced 18-[13C]-oleic acid from at least 3 known fatty acid binding sites on HSA that also bind many drugs. Binding isotherms of testosterone's binding to HSA generated using fluorescence spectroscopy and equilibrium dialysis were nonlinear and the apparent dissociation constant varied with different concentrations of testosterone and HSA. The binding isotherms neither conformed to a linear binding model with 1:1 stoichiometry nor to 2 independent binding sites; the binding isotherms were most consistent with 2 or more allosterically coupled binding sites. Molecular dynamics studies revealed that testosterone's binding to fatty acid binding site 3 on HSA was associated with conformational changes at site 6, indicating that residues in in these 2 distinct binding sites are allosterically coupled. There are multiple, allosterically coupled binding sites for testosterone on HSA. Testosterone shares these binding sites on HSA with free fatty acids, which could displace testosterone from HSA under various physiological states or disease conditions, affecting its bioavailability.

Development of a theoretical model for the inhibition of nsP3 protease of Chikungunya virus using pyranooxazoles.

Chikungunya virus (CHIKV) causes Chikungunya fever (CHIKF) and till date no effective medicine for its cure is available in market. Different research groups find various possible interactions between small molecules and non-structural proteins, viz. nsP3, one of the most important viral elements in CHIKV. In this work, authors have studied the interactions of nsP3 protease of CHIKV with pyranooxazoles. Initially, a one-pot three-component reaction was designed using oxazolidine-2,4-dione, benzaldehyde and cyanoethylacetate to get a proposed biological active molecule, i.e. based on pyranooxazoles. The mechanism for the synthesis of the product based on pyranooxazole was studied through density functional theory (DFT) using Gaussian. Then, a library of the obtained pyranooxazole was created through computational tools by varying the substituents. Further, virtual screening of the designed library of pyranooxazoles (200 compounds) against nsP3 protease of CHIKV was performed. Herein, CMPD 104 showed strongest binding affinity toward the targeted nsP3 protease of CHIKV, based on the least binding energy obtained from docking. Based on docking results, the pharmacological, toxicity, biological score and Lipinski's filters were studied. Further, DFT studies of top five compounds were done using Gaussian. Molecular dynamics (MD) simulation of nsP3 protease of CHIKV with and without 104 was performed using AMBER18 utilizing ff14SB force field in three steps (minimization, equilibration and production). This work is emphasized to designing of one-pot three-component synthesis and to develop a theoretical model to inhibit the nsP3 protease of CHIKV. AbbreviationsCHIKFChikungunya feverCHIKVChikungunya virusDFTdensity functional theoryDSDiscovery StudioMDmolecular dynamicsMM-GBSAmolecular mechanics-generalized born surface areaMMVMolegro molecular viewerCommunicated by Ramaswamy H. Sarma.

In-silico prediction of novel drug-target complex of nsp3 of CHIKV through molecular dynamic simulation.

Literature reported that nsp3 of CHIKV is an important target for the designing of drug as it involves in the replication, survival etc. Herein, about eighteen million molecules available in the ZINC database are filtered against nsp3 using RASPD. Top five hit drug molecules were then taken from the total screened molecules (6988) from ZINC database. Then, a one pot-three components reaction is designed to get the pyrazolophthalazine and its formation was studied using DFT method. Authors created a library of 200 compounds using the product obtained in the reaction and filtered against nsp3 of CHIKV based on docking using iGEMDOCK, a computational tool. Authors have studied the best molecules after applying the the Lipinski's rule of five and bioactive score. Further, the authors took the best compound i.e. CMPD178 and performed the MD simulations and tdMD simulations with nsp3 protease using AMBER18. MD trajectories were studied to collect the information about the nsp3 of CHIKV with and without screened compound and then, MM-GBSA calculations were performed to calculate change in binding free energies for the formation of complex. The aim of the work is to find the potential candidate as promising inhibitor against nsp3 of CHIKV.

Computational approach to study the synthesis of noscapine and potential of stereoisomers against nsP3 protease of CHIKV.

Chikungunya fever is a major public health issue in India affecting millions of people and occurs due to Chikungunya. Chikungunya virus (CHIKV) is a single stranded RNA virus from the family of Togaviridae and genus alpha virus. It contain three structural proteins: glycosylated E1 and E2, embedded in the viral envelope, and a non-glycosylated nucleocapsid protein. Till date, researchers are working on inhibition of CHIKV but till now no cheap and effective medicine is available in the market. Therefore, the authors of this work thought of isoquinoline based noscapine to inhibit the nsP3 protease of CHIKV. The aim of the work is to understand the mechanism for the synthesis of noscapine theoretically using DFT. Further study the potential of all four isomers of noscapines {(13 (S,R), 14 (R,R), 15 (R,S) and 16 (S,S)} against nsP3 protease of CHIKV with the help of docking and MD simulation. The integrated e-pharmacophore binding affinity based virtual screening, docking and molecular dynamics simulation recognized four hits isomers as inhibition nsP3 protease of CHIKV. The docking energies of all the isomers of noscapine (13-16) with nsP3 protease CHIKV was found out to be more negative than baicalin (-8.06 kcal/mol) on selected sites. Amongst the isomers of noscapine, CMPD 13 possessed best binding affinity with four hydrogen bonding interactions. Further, ADME properties and blood-brain barrier permeability properties have been calculated. DFT studies of all the isomers of noscapine was investigated.

Trimethylamine-N-oxide switches from stabilizing nature: A mechanistic outlook through experimental techniques and molecular dynamics simulation.

In adaptation biology of the discovery of the intracellular osmolytes, the osmolytes are found to play a central role in cellular homeostasis and stress response. A number of models using these molecules are now poised to address a wide range of problems in biology. Here, a combination of biophysical measurements and molecular dynamics (MD) simulation method is used to examine the effect of trimethylamine-N-oxide (TMAO) on stem bromelain (BM) structure, stability and function. From the analysis of our results, we found that TMAO destabilizes BM hydrophobic pockets and active site as a result of concerted polar and non-polar interactions which is strongly evidenced by MD simulation carried out for 250 ns. This destabilization is enthalpically favourable at higher concentrations of TMAO while entropically unfavourable. However, to the best of our knowledge, the results constitute first detailed unambiguous proof of destabilizing effect of most commonly addressed TMAO on the interactions governing stability of BM and present plausible mechanism of protein unfolding by TMAO.