Abstract
Small molecule inhibitors of snake venom metalloproteinases (SVMPs) could provide a means to rapidly halt the progression of local tissue damage following viperid snake envenomations. In this study, we examine the ability of candidate compounds based on a pentacyclic triterpene skeleton to inhibit SVMPs. We leverage molecular dynamics simulations to estimate the free energies of the candidate compounds for binding to BaP1, a P-I type SVMP, and compare these results with experimental assays of proteolytic activity inhibition in a homologous enzyme (Batx-I). Both simulation and experiment suggest that betulinic acid is the most active candidate, with the simulations predicting a standard binding free energy of kcal/mol. The simulations also reveal the atomic interactions that underlie binding between the triterpenic acids and BaP1, most notably the electrostatic interaction between carboxylate groups of the compounds and the zinc cofactor of BaP1. Together, our simulations and experiments suggest that occlusion of the S1 subsite is essential for inhibition of proteolytic activity. While all active compounds make hydrophobic contacts in the S1 site, -boswellic acid, with its distinct carboxylate position, does not occlude the S1 site in simulation and exhibits negligible activity in experiment.
Highlights
Snake venom metalloproteinases (SVMPs) are zinc-dependent hydrolases that represent a major component in most viperid venoms and are classified into groups PI to PIII according to their domain organization [1]
The simulations demonstrate that binding of these compounds is driven by a strong electrostatic interaction between their carboxylate group and the catalytic Zn2+ ion of the metalloproteinase, which is further reinforced by hydrophobic interactions of the A, B and C rings
Explicit-solvent molecular dynamic simulations coupled with the adaptive biasing force (ABF)
Summary
Snake venom metalloproteinases (SVMPs) are zinc-dependent hydrolases that represent a major component in most viperid venoms and are classified into groups PI to PIII according to their domain organization [1]. Catalytic cleavage in SVMPs is mediated by a Zn2+ ion coordinated by three conserved histidine side chains, and a water molecule anchored to a glutamate residue. Following the common nomenclature for peptide/protease complexes, the metalloproteinase substrate binding site is divided into several subsites (regions on the enzyme surface that interact with individual amino acid residues on either side of the substrate cleavage site) [3]. Toxins 2018, 10, 397 the cleavage region are labeled as S1, S2, S3 (unprimed subsites), while those on the carboxyl side are labeled as S10 , S20 , S30 (primed subsites).
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