A computational H5N1 neuraminidase model and its binding to commercial drugs

In order to understand the mechanisms of ligand binding and interaction between two commercial drugs (ligands), zanamivir and oseltamivir and H5N1 Influenza Virus Neuraminidase subtype N1, a three-dimensional model of N1-ligand (GenBank accession no. AAS654617) was initially generated by homology modeling using the 13 high-resolution X-ray structures of neuraminidase N2 and N9 as the template. With the aid of the molecular mechanics and molecular dynamics methods, the final implicit solvent refined model was obtained. It was, then, assessed by PROCHECK, PROSA and VERIFY3D. With this model, a flexible docking study was performed. The results show strong hydrogen bond interactions between the glycerol side chains of zanamivir and Arg29 of the N1. Common hydrogen bonds between the carboxyl groups and Arg279 were found for both drugs. It was also found that the Glu30, Asp62, Arg63, Arg204, Trp310, Tyr313, Glu336, Ile338, Trp348, Ala349 were observed to facilitate the enzyme-ligand non-bonding interactions as they are located within the radius of 5 Å from all atoms of both drugs. Charge distribution was evaluated using the semi-empirical AM1 method. The results show that the total net charges of the –NH side chain of zanamivir is less negative than that of oseltamivir. This is in contrast to what is observed for the amide and alkyl (ether/glycerol) side chains. In comparison of the binding free energies between the X-ray N2-ligand and N9-ligand complexes, N1-ligand binding is found to be less potent than N2 and N9 subtypes, while N2-ligand and N9-ligand are roughly comparable. In addition, it is interesting to observe that the binding free energies for all three subtypes of the zanamivir complexes are lower than those of oseltamivir.     

Quantum mechanics/molecular mechanics modeling of substrate-assisted catalysis in family 18 chitinases: conformational changes and the role of Asp142 in catalysis in ChiB

Family 18 chitinases catalyze the hydrolysis of β-1,4-glycosidic bonds in chitin. The mechanism has been proposed to involve the formation of an oxazolinium ion intermediate via an unusual substrate-assisted mechanism, in which the substrate itself acts as an intramolecular nucleophile (instead of an enzyme residue). Here, we have modeled the first step of the chitin hydrolysis catalyzed by Serratia marcescens chitinase B for the first time using a combined quantum mechanics/molecular mechanics approach. The calculated reaction barriers based on multiple snapshots are 15.8-19.8 kcal mol(-1) [B3LYP/6-31+G(d)//AM1-CHARMM22], in good agreement with the activation free energy of 16.1 kcal mol(-1) derived from experiment. The enzyme significantly stabilizes the oxazolinium intermediate. Two stable conformations ((4)C(1)-chair and B(3,O)-boat) of the oxazolinium ion intermediate in subsite -1 were unexpectedly observed. The transition state structure has significant oxacarbenium ion-like character. The glycosyl residue in subsite -1 was found to follow a complex conformational pathway during the reaction ((1,4)B → [(4)H(5)/(4)E](++) → (4)C(1) ? B(3,O)), indicating complex conformational behavior in glycoside hydrolases that utilize a substrate-assisted catalytic mechanism. The D142N mutant is found to follow the same wild-type-like mechanism: the calculated barriers for reaction in this mutant (16.0-21.1 kcal mol(-1)) are higher than in the wild type, in agreement with the experiment. Asp142 is found to be important in transition state and intermediate stabilization.     

Theoretical study of the arabinan hydrolysis by an inverting GH43 arabinanase

The endo-1,5-α-L-arabinanases (ABNases) belong to the glycoside hydrolase family 43 (GH43) and hydrolyse α-1,5-arabinofuranosidic bonds in arabinose-containing polysaccharides by an inverting mechanism. ABNases are key enzymes involved in hemicellulose degradation with various applications in food technology, organic synthesis and biofuel production. Here, the reaction mechanism of the Geobacillus stearothermophilus GH43 ABNases has been investigated using density functional theory method. To probe the role of the pKa modulators (Ser164 and Tyr229) in the reaction, two different orientations of a catalytic acid (Glu201) with respect to the position of these residues are considered. The results show that the orientation involving a hydrogen bond network (Ser164???Glu201???Tyr229) is energetically more favourable, compared to the other one (Ser164???Glu201), with a barrier of 13.6 kcal mol^?1, consistent with the experimental data. Our calculations also give support to a single displacement mechanism proposed in the literature where the protonation of glycosidic bond and a nucleophilic attack by a catalytic water occur in a concerted but asynchronous manner.     

Inhibitory Potential of a Designed Peptide Inhibitor Based on Zymogen Structure of Trypsin from Spodoptera frugiperda: In Silico Insights

Spodoptera frugiperda (J.E. Smith) is an invasive pest in agriculture. It can potentially damage yield resulting severe crop losses and subsequently significant economic damage each year. S. frugiperda is predominantly managed using traditional chemical pesticides. Accordingly, sustainable alternatives such as digestive enzymes inhibitors can be used as an efficient pest management that protects the environment. This contribution aims to examine the pro-region of S. frugiperda trypsin as specific inhibitor of the pest protease enzyme. Structural modeling in conjunction with molecular docking simulations were conducted to design a peptide sequence with the best docking scores and strong binding energy to the target enzyme. The structural models of six pro-peptides were produced based on modification of 7-amino acids of the pro-region of S. frugiperda trypsin. VERIFY_3D, ERRAT, PROCHECK, PROSA and WHAT-IF scores validated the reliability of the predicted model of S. frugiperda trypsin. Molecular docking studies between the six designed inhibitor peptides and the predicted model structure at three different pH conditions were carried out. Data revealed that VPSNPQR at pH 11.0 with the best docking score, the lowest binding energy (ΔG) and dissociation constant (K_d) indicated a potent binding affinity towards S. frugiperda trypsin’s active site. Moreover, the peptide showed a weak potential for interaction with the human trypsin. The results indicated the importance of computational studies in design and selection of inhibitor peptides against target enzymes. Such inhibitors can be used for S. frugiperda control, which can be further applied in other pest management programs.

Graphical Abstract

Docking simulations between the pro-peptide inhibitor and Spodoptera frugiperda midgut trypsin confirmed the capacity of the designed pro-region in inhibiting the insect trypsin.