Protein-substrate recognition is highly dynamic and complex process in nature. A key approach in deciphering the mechanism underlying the recognition process is to capture the kinetic process of substrate in its act of binding to its designated protein cavity. Towards this end, microsecond long atomistic molecular dynamics (MD) simulation has recently emerged as a popular method of choice, due its ability to record these events at high spatial and temporal resolution. However, success in this approach comes at an exorbitant computational cost. Here we demonstrate that coarse grained models of protein, when systematically optimised to maintain its tertiary fold, can capture the complete process of spontaneous protein-ligand binding from bulk media to cavity, within orders of magnitude shorter wall clock time compared to that of all-atom MD simulations. The simulated and crystallographic binding pose are in excellent agreement. We find that the exhaustive sampling of ligand exploration in protein and solvent, harnessed by coarse-grained simulation at a frugal computational cost, in combination with Markov state modelling, leads to clearer mechanistic insights and discovery of novel recognition pathways. The result is successfully validated against three popular protein-ligand systems. Overall, the approach provides an affordable and 1 . CC-BY-NC-ND 4.

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Achieving superfast water transport by synthetically designed molecular artifacts, which exclude salts and protons, is a challenging task in separation science today, as it requires the concomitant presence of a...

Amyloid aggregation of protein is linked to many neurodegenerative diseases. Identification of small molecules capable of targeting amyloidogenic proteins has gained significant importance. Introduction of hydrophobic and hydrogen bonding interactions through site-specific binding of small molecular ligand to protein can effectively modulate the protein aggregation pathway. Here, we investigate the possible roles of three different bile acids, cholic acid (CA), taurocholic acid (TCA), and lithocholic acid (LCA) with varying hydrophobic and hydrogen bonding properties in inhibiting protein fibrillation. Bile acids are an important class of steroid compounds that are synthesized in the liver from cholesterol. Increasing evidence suggests that altered taurine transport, cholesterol metabolism, and bile acid synthesis have strong implications in Alzheimer’s disease. We find that the hydrophilic bile acids, CA and TCA (taurine conjugated form of CA), are substantially more efficient inhibitors of lysozyme fibrillation than the most hydrophobic secondary bile acid LCA. Although LCA binds more strongly with the protein and masks the Trp residues more prominently through hydrophobic interactions, the lesser extent of hydrogen bonding interactions at the active site has made LCA a relatively weaker inhibitor of HEWL aggregation than CA and TCA. The introduction of a greater number of hydrogen bonding channels by CA and TCA with several key amino acid residues which are prone to form oligomers and fibrils has weakened the protein’s internal hydrogen bonding capabilities for undergoing amyloid aggregation.

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Cytochrome c (Cyt-c) is a heme-containing protein that plays a vital role as an electron carrier in the mitochondrial respiratory chain. Cyt-c possesses peroxidase-like activity in the native state despite its six-coordinated heme iron. In this work, we studied the effect of magnetic ionic liquids (MILs) on the peroxidase activity of Cyt-c. Guaiacol oxidation was used to probe the activity of Cyt-c in the presence of cholinium (Ch)-based MILs such as [Ch][FeCl4] and [Ch]2[MnCl4]. A detailed kinetics and thermodynamics study on the activity of Cyt-c in the presence of MILs was investigated with the help of isothermal titration calorimetry (ITC), UV–vis, circular dichroism (CD), and fluorescence spectroscopy. The peroxidase activity of Cyt-c increases by twofold in the presence of [Ch][FeCl4] and only 20% in the presence of [Ch]2[MnCl4] IL. This correlates with the accessibility of the heme iron of Cyt-c. Molecular docking and molecular dynamic simulations were also employed to explore the mechanism of the interaction. The increase in the peroxidase activity of Cyt-c is attributed to the perturbation in the native sixth coordination bond of methionine-80 associated with the heme region of Cyt-c. Since not much work has been done on the enzymatic activity in the presence of MILs, we hope this will set the platform to use MILs to modulate and control enzymatic activity and catalysis.

Ras GTPase is an enzyme that catalyzes the hydrolysis of guanosine triphosphate (GTP) and plays an important role in controlling crucial cellular signaling pathways. However, this enzyme has always been believed to be undruggable due to its strong binding affinity with its native substrate GTP. To understand the potential origin of high GTPase/GTP recognition, here we reconstruct the complete process of GTP binding to Ras GTPase via building Markov state models (MSMs) using a 0.1 ms long all-atom molecular dynamics (MD) simulation. The kinetic network model, derived from the MSM, identifies multiple pathways of GTP en route to its binding pocket. While the substrate stalls onto a set of non-native metastable GTPase/GTP encounter complexes, the MSM accurately discovers the native pose of GTP at its designated catalytic site in crystallographic precision. However, the series of events exhibit signatures of conformational plasticity in which the protein remains trapped in multiple non-native conformations even when GTP has already located itself in its native binding site. The investigation demonstrates mechanistic relays pertaining to simultaneous fluctuations of switch 1 and switch 2 residues which remain most instrumental in maneuvering the GTP-binding process. Scanning of the crystallographic database reveals close resemblance between observed non-native GTP binding poses and precedent crystal structures of substrate-bound GTPase, suggesting potential roles of these binding-competent intermediates in allosteric regulation of the recognition process.

Protein-substrate recognition is highly dynamic and complex process in nature. A key approach in deciphering the mechanism underlying the recognition process is to capture the kinetic process of substrate in its act of binding to its designated protein cavity. Towards this end, microsecond long atomistic molecular dynamics (MD) simulation has recently emerged as a popular method of choice, due its ability to record these events at high spatial and temporal resolution. However, success in this approach comes at an exorbitant computational cost. Here we demonstrate that coarse grained models of protein, when systematically optimised to maintain its tertiary fold, can capture the complete process of spontaneous protein-ligand binding from bulk media to cavity, within orders of magnitude shorter wall clock time compared to that of all-atom MD simulations. The simulated and crystallographic binding pose are in excellent agreement. We find that the exhaustive sampling of ligand exploration in protein and solvent, harnessed by coarse-grained simulation at a frugal computational cost, in combination with Markov state modelling, leads to clearer mechanistic insights and discovery of novel recognition pathways. The result is successfully validated against three popular protein-ligand systems. Overall, the approach provides an affordable and 1 attractive alternative of all-atom simulation and promises a way-forward for replacing traditional docking based small molecule discovery by high-throughput coarse-grained simulation for searching potential binding site and allosteric sites. This also provides practical avenues for first-hand exploration of bio-molecular recognition processes in large-scale biological systems, otherwise inaccessible in all-atom simulations.