Snake venom metalloproteinases
(SVMPs) are important
drug targets
against snakebite envenoming, the neglected tropical disease with
the highest mortality worldwide. Here, we focus on Russell’s
viper (Daboia russelii), one of the
“big four” snakes of the Indian subcontinent that, together,
are responsible for ca. 50,000 fatalities annually. The “Russell’s
viper venom factor X activator” (RVV-X), a highly toxic metalloproteinase,
activates the blood coagulation factor X (FX), leading to the prey’s
abnormal blood clotting and death. Given its tremendous public health
impact, the WHO recognized an urgent need to develop efficient, heat-stable,
and affordable-for-all small-molecule inhibitors, for which a deep
understanding of the mechanisms of action of snake’s principal
toxins is fundamental. In this study, we determine the catalytic mechanism
of RVV-X by using a density functional theory/molecular mechanics
(DFT:MM) methodology to calculate its free energy profile. The results
showed that the catalytic process takes place via two steps. The first
step involves a nucleophilic attack by an in situ generated hydroxide
ion on the substrate carbonyl, yielding an activation barrier of 17.7
kcal·mol–1, while the second step corresponds
to protonation of the peptide nitrogen and peptide bond cleavage with
an energy barrier of 23.1 kcal·mol–1. Our study
shows a unique role played by Zn2+in catalysis by lowering
the pKaof the Zn2+-bound water
molecule, enough to permit the swift formation of the hydroxide nucleophile
through barrierless deprotonation by the formally much less basic
Glu140. Without the Zn2+cofactor, this step would be rate-limiting.