scholarly journals Essential Loop Dynamics Modulates Catalytic Activity in α-Chymotrypsin

2021 ◽  
Author(s):  
Pritam Biswas ◽  
Uttam Pal ◽  
Aniruddha Adhikari ◽  
Susmita Mondal ◽  
Ria Ghosh ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Md Simulation ◽  
Conformational Dynamics ◽  
Principal Component ◽  
Catalytic Efficiency ◽  
Md Simulations ◽  
Essential Dynamics ◽  
Loop Dynamics ◽  
Loop Regions

Conformational dynamics of macromolecules including enzymes are essential for their function. The present work reports the role of essential dynamics in alpha-chymotrypsin (CHT) which correlates with its catalytic activity. Detailed optical spectroscopy and classical molecular dynamics (MD) simulation were used to study thermal stability, catalytic activity and dynamical flexibility of the enzyme. The study of the enzyme kinetics reveals an optimum catalytic efficiency at 308K. Polarization gated fluorescence anisotropy with 8-anilino-1-napthelene sulfonate (ANS) have indicated increasing flexibility of the enzyme with an increase in temperature. Examination of the structure of CHT reveal the presence of five loop regions (LRs) around the catalytic S1 pocket. MD simulations have indicated that flexibility increases concurrently with temperature which decreases beyond optimum temperature. Principal component analysis (PCA) of the eigenvectors manifests essential dynamics and gatekeeping role of the five LRs surrounding the catalytic pocket which controls the enzyme activity.

CLUSTERING MULTI-DOMAIN PROTEIN STRUCTURES IN THE ESSENTIAL DYNAMICS SUBSPACE

2013 ◽  
Vol 12 (08) ◽  
pp. 1341008 ◽  
Author(s):  
BIN WEN ◽  
YUNYU SHI ◽  
ZHIYONG ZHANG
Keyword(s):  
Cluster Analysis ◽  
Md Simulation ◽  
Clustering Algorithm ◽  
Protein Structures ◽  
Principal Component ◽  
Md Simulations ◽  
Essential Dynamics ◽  
Ww Domains ◽  
Domain Protein ◽  
Domain Motions

A multi-domain protein is able to exist as equilibrium of different conformations in solution, which may be critical to its biological function. Besides experimental techniques, computational methods like molecular dynamics (MD) simulations are suitable to study inter-domain motions of the protein and sample different conformational states. A MD simulation usually generates a trajectory containing large amount of protein structures, and a post-processing cluster analysis would be necessary to group similar structures into clusters and identify these typical conformations of the multi-domain protein. In this paper, the widely used k-means clustering algorithm is implemented in the protein essential dynamics (ED) subspace defined by principal component analysis on the MD trajectory. Cluster analysis of the formin binding protein 21 (FBP21) tandem WW domains demonstrate that the k-means clustering results by measuring distances between structures in the ED subspace are superior to those by using other metrics like pairwise inter-domain residue distances.

scholarly journals Structure of the full-length human Pannexin1 channel and insights into its role in pyroptosis

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Sensen Zhang ◽  
Baolei Yuan ◽  
Jordy Homing Lam ◽  
Jun Zhou ◽  
Xuan Zhou ◽  
...  
Keyword(s):  
Md Simulation ◽  
Potential Role ◽  
Md Simulations ◽  
Full Length ◽  
Inflammasome Activation ◽  
Simulation Studies ◽  
Cryo Electron Microscopy ◽  
Gating Mechanism ◽  
Atp Efflux

AbstractPannexin1 (PANX1) is a large-pore ATP efflux channel with a broad distribution, which allows the exchange of molecules and ions smaller than 1 kDa between the cytoplasm and extracellular space. In this study, we show that in human macrophages PANX1 expression is upregulated by diverse stimuli that promote pyroptosis, which is reminiscent of the previously reported lipopolysaccharide-induced upregulation of PANX1 during inflammasome activation. To further elucidate the function of PANX1, we propose the full-length human Pannexin1 (hPANX1) model through cryo-electron microscopy (cryo-EM) and molecular dynamics (MD) simulation studies, establishing hPANX1 as a homo-heptamer and revealing that both the N-termini and C-termini protrude deeply into the channel pore funnel. MD simulations also elucidate key energetic features governing the channel that lay a foundation to understand the channel gating mechanism. Structural analyses, functional characterizations, and computational studies support the current hPANX1-MD model, suggesting the potential role of hPANX1 in pyroptosis during immune responses.

scholarly journals Tethering-induced destabilization and ATP-binding for tandem RRM domains of ALS-causing TDP-43 and hnRNPA1

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Mei Dang ◽  
Yifan Li ◽  
Jianxing Song
Keyword(s):  
Conformational Dynamics ◽  
Md Simulations ◽  
Atp Binding ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Nucleic Acid Binding ◽  
General Role ◽  
Nmr Characterization ◽  
Lateral Sclerosis

AbstractTDP-43 and hnRNPA1 contain tandemly-tethered RNA-recognition-motif (RRM) domains, which not only functionally bind an array of nucleic acids, but also participate in aggregation/fibrillation, a pathological hallmark of various human diseases including amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), alzheimer's disease (AD) and Multisystem proteinopathy (MSP). Here, by DSF, NMR and MD simulations we systematically characterized stability, ATP-binding and conformational dynamics of TDP-43 and hnRNPA1 RRM domains in both tethered and isolated forms. The results reveal three key findings: (1) upon tethering TDP-43 RRM domains become dramatically coupled and destabilized with Tm reduced to only 49 °C. (2) ATP specifically binds TDP-43 and hnRNPA1 RRM domains, in which ATP occupies the similar pockets within the conserved nucleic-acid-binding surfaces, with the affinity slightly higher to the tethered than isolated forms. (3) MD simulations indicate that the tethered RRM domains of TDP-43 and hnRNPA1 have higher conformational dynamics than the isolated forms. Two RRM domains become coupled as shown by NMR characterization and analysis of inter-domain correlation motions. The study explains the long-standing puzzle that the tethered TDP-43 RRM1–RRM2 is particularly prone to aggregation/fibrillation, and underscores the general role of ATP in inhibiting aggregation/fibrillation of RRM-containing proteins. The results also rationalize the observation that the risk of aggregation-causing diseases increases with aging.

scholarly journals Computational Studies of SARS-CoV-2 3CLpro: Insights from MD Simulations

2020 ◽  
Vol 21 (15) ◽  
pp. 5346 ◽  
Author(s):  
Alessandro Grottesi ◽  
Neva Bešker ◽  
Andrew Emerson ◽  
Candida Manelfi ◽  
Andrea R. Beccari ◽  
...  
Keyword(s):  
Conformational Dynamics ◽  
Md Simulations ◽  
Health Impact ◽  
Respiratory Syndrome Virus ◽  
Enzymatic Function ◽  
Main Protease ◽  
Loop Regions ◽  
Dimeric Form ◽  
And Function ◽  
Shed Light

Given the enormous social and health impact of the pandemic triggered by severe acute respiratory syndrome 2 (SARS-CoV-2), the scientific community made a huge effort to provide an immediate response to the challenges posed by Coronavirus disease 2019 (COVID-19). One of the most important proteins of the virus is an enzyme, called 3CLpro or main protease, already identified as an important pharmacological target also in SARS and Middle East respiratory syndrome virus (MERS) viruses. This protein triggers the production of a whole series of enzymes necessary for the virus to carry out its replicating and infectious activities. Therefore, it is crucial to gain a deeper understanding of 3CLpro structure and function in order to effectively target this enzyme. All-atoms molecular dynamics (MD) simulations were performed to examine the different conformational behaviors of the monomeric and dimeric form of SARS-CoV-2 3CLpro apo structure, as revealed by microsecond time scale MD simulations. Our results also shed light on the conformational dynamics of the loop regions at the entry of the catalytic site. Studying, at atomic level, the characteristics of the active site and obtaining information on how the protein can interact with its substrates will allow the design of molecules able to block the enzymatic function crucial for the virus.

scholarly journals Stabilization and ATP-binding for tandem RRM domains of ALS-causing TDP-43 and hnRNPA1

2020 ◽  
Author(s):  
Mei Dang ◽  
Yifan Li ◽  
Jianxing Song
Keyword(s):  
Nucleic Acid ◽  
Nucleic Acids ◽  
Conformational Dynamics ◽  
Md Simulations ◽  
Human Diseases ◽  
Atp Binding ◽  
Nucleic Acid Binding ◽  
General Role ◽  
Nmr Characterization

AbstractTDP-43 and hnRNPA1 contain tandemly-tethered RRM domains, which not only functionally bind an array of nucleic acids, but also participate in aggregation/fibrillation, a pathological hallmark of various human diseases including ALS, FTD, AD and MSP. Here, by DSF, NMR and MD simulations we systematically characterized stability, ATP-binding and conformational dynamics of TDP-43 and hnRNPA1 RRM domains in both tethered and isolated forms. The results reveal three key findings: 1) very unexpectedly, upon tethering TDP-43 RRM domains become dramatically coupled and destabilized with Tm reduced to only 49 °C. 2) ATP specifically binds TDP-43 and hnRNPA1 RRM domains, in which ATP occupies the similar pockets within the conserved nucleic-acid-binding surfaces, with the affinity higher to the tethered than isolated forms. 3) MD simulations indicate that the tethered RRM domains of TDP-43 and hnRNPA1 have higher conformational dynamics than the isolated forms. Two RRM domains become coupled as shown by NMR characterization and analysis of inter-domain correlation motions. The study explains the long-standing puzzle that the tethered TDP-43 RRM1-RRM2 is particularly prone to aggregation/fibrillation, and underscores the general role of ATP in inhibiting aggregation/fibrillation of RRM-containing proteins. The results also rationalize the observation that the risk of aggregation-causing diseases increases with aging.

scholarly journals Kinetic Studies on CphA Mutants Reveal the Role of the P158-P172 Loop in Activity versus Carbapenems

2016 ◽  
Vol 60 (5) ◽  
pp. 3123-3126 ◽  
Author(s):  
Carlo Bottoni ◽  
Mariagrazia Perilli ◽  
Francesca Marcoccia ◽  
Alessandra Piccirilli ◽  
Cristina Pellegrini ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Catalytic Efficiency ◽  
Kinetic Studies ◽  
Zinc Ion ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Wild Type ◽  
The Stability

ABSTRACTSite-directed mutagenesis of CphA indicated that prolines in the P158-P172 loop are essential for the stability and the catalytic activity of subclass B2 metallo-β-lactamases against carbapenems. The sequential substitution of proline led to a decrease of the catalytic efficiency of the variant compared to the wild-type (WT) enzyme but also to a higher affinity for the binding of the second zinc ion.

scholarly journals P174E Substitution in GES-1 and GES-5 β-Lactamases Improves Catalytic Efficiency toward Carbapenems

2018 ◽  
Vol 62 (5) ◽  
Author(s):  
Alessandra Piccirilli ◽  
Paola Sandra Mercuri ◽  
Moreno Galleni ◽  
Massimiliano Aschi ◽  
André Matagne ◽  
...  
Keyword(s):  
Clavulanic Acid ◽  
Catalytic Efficiency ◽  
Md Simulations ◽  
Hydrolytic Activity ◽  
Cd Spectroscopy ◽  
Saturation Mutagenesis ◽  
Wild Type ◽  
Structural Content ◽  
Drastic Increase

ABSTRACT GES-type β-lactamases are a group of enzymes that have evolved their hydrolytic activity against carbapenems. In this study, the role of residue 174 inside the Ω-loop of GES-1 and GES-5 was investigated. GES-1 P174E and GES-5 P174E mutants, selected by site saturation mutagenesis, were purified and kinetically characterized. In comparison with GES-1 and GES-5 wild-type enzymes, GES-1 P174E and GES-5 P174E mutants exhibited lower k cat and k cat / K m values for cephalosporins and penicillins. Concerning carbapenems, GES-1 P174E shared higher k cat values but lower K m values than those calculated for GES-1. The GES-1 P174E and GES-5 P174E mutants showed high hydrolytic efficiency for imipenem, with k cat / K m values 100- and 660-fold higher, respectively, than those of GES-1. Clavulanic acid and tazobactam are good inhibitors for both GES-1 P174E and GES-5 P174E . Molecular dynamic (MD) simulations carried out for GES-1, GES-5, GES-1 P174E , and GES-5 P174E complexed with imipenem and meropenem have shown that mutation at position 174 induces a drastic increase of enzyme flexibility, in particular in the Ω-loop. The circular dichroism (CD) spectroscopy spectra of the four enzymes indicate that the P174E substitution in GES-1 and GES-5 does not affect the secondary structural content of the enzymes.

Using molecular simulation to explore the nanoscale dynamics of the plant kinome

2018 ◽  
Vol 475 (5) ◽  
pp. 905-921 ◽  
Author(s):  
Alexander S. Moffett ◽  
Diwakar Shukla
Keyword(s):  
Molecular Simulation ◽  
Conformational Changes ◽  
Cellular Response ◽  
Md Simulation ◽  
Conformational Dynamics ◽  
Large Family ◽  
Md Simulations ◽  
The Future ◽  
Catalytically Active ◽  
Domains Of Life

Eukaryotic protein kinases (PKs) are a large family of proteins critical for cellular response to external signals, acting as molecular switches. PKs propagate biochemical signals by catalyzing phosphorylation of other proteins, including other PKs, which can undergo conformational changes upon phosphorylation and catalyze further phosphorylations. Although PKs have been studied thoroughly across the domains of life, the structures of these proteins are sparsely understood in numerous groups of organisms, including plants. In addition to efforts towards determining crystal structures of PKs, research on human PKs has incorporated molecular dynamics (MD) simulations to study the conformational dynamics underlying the switching of PK function. This approach of experimental structural biology coupled with computational biophysics has led to improved understanding of how PKs become catalytically active and why mutations cause pathological PK behavior, at spatial and temporal resolutions inaccessible to current experimental methods alone. In this review, we argue for the value of applying MD simulation to plant PKs. We review the basics of MD simulation methodology, the successes achieved through MD simulation in animal PKs, and current work on plant PKs using MD simulation. We conclude with a discussion of the future of MD simulations and plant PKs, arguing for the importance of molecular simulation in the future of plant PK research.

scholarly journals Role of the Outer Metal of Double Metal Cyanides on the Catalytic Efficiency in Styrene Oxidation

Catalysts ◽  
2019 ◽  
Vol 9 (11) ◽  
pp. 905
Author(s):  
Paulina Molina-Maldonado ◽  
Rosario Ruíz-Guerrero ◽  
Carlos Hernández-Fuentes
Keyword(s):  
Catalytic Activity ◽  
Divalent Cations ◽  
Catalytic Efficiency ◽  
Oxidizing Agent ◽  
Styrene Oxidation ◽  
Metal Cyanide ◽  
Metal Cyanides ◽  
Model Molecule ◽  
Tert Butyl Hydroperoxide

The catalytic efficiency of double metal cyanide (DMC) has been shown to be very effective in heterogeneous catalysis. The catalytic activity of the outer divalent cations (Mn, Co, Ni, and Cu) of a family of hexacyanocobaltates was examined in the oxidation reaction of styrene, as a model molecule, using tert Butyl Hydroperoxide (TBHP, Luperox®) as an oxidizing agent. The most electronegative outer cations showed the best conversions, with 95% for copper, followed by nickel with 85% conversion of the monomer at atmospheric pressure and temperature of 75 °C. The evidence showed that the catalytic activity and selectivity towards oxidized products are strongly linked to the accurate choice of the outer cation in the DMC together with the oxidizing agent.

scholarly journals Investigating the conformational dynamics of SARS-CoV-2 NSP6 protein with emphasis on non-transmembrane 91-112 & 231-290 regions

2021 ◽  
Author(s):  
Amit Kumar ◽  
Prateek Kumar ◽  
Kumar Udit Saumya ◽  
Rajanish Giri
Keyword(s):  
Structural Information ◽  
Transmembrane Protein ◽  
Conformational Dynamics ◽  
Md Simulations ◽  
Host Protein ◽  
Helical Conformation ◽  
Beta Sheet ◽  
Host Proteins ◽  
Helical Structures

The NSP6 protein of SARS-CoV-2 is a transmembrane protein, with some regions lying outside the membrane. Besides, a brief role of NSP6 in autophagosome formation, this is not studied significantly. Also, there is no structural information available till date. Based on the prediction by TMHMM server for transmembrane prediction, it is found that the N-terminal residues (1-11), middle region residues (91-112) and C-terminal residues (231-290) lies outside the membrane. Molecular Dynamics (MD) simulations showed that NSP6 consisting of helical structures, whereas membrane outside lying region (91-112) showed partial helicity, which further used as model and obtain disordered type conformation after 1.5 microseconds. Whereas, the residues 231-290 has both helical and beta sheet conformations in its structure model. A 200ns simulations resulted in the loss of beta sheet structures, while helical regions remained intact. Further, we have characterized the residue 91-112 by using reductionist approaches. The NSP6 (91-112) was found disordered like in isolation, which gain helical conformation in different biological mimic environmental conditions. These studies can be helpful to study NSP6 (91-112) interactions with host proteins, where different protein conformation might play significant role. The present study adds up more information about NSP6 protein aspect, which could be exploited for its host protein interaction and pathogenesis.

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