Molecular binding scaffolds increase local substrate concentration enhancing the enzymatic hydrolysis of VX nerve agent

Abstract

Kinetic enhancement of organophosphate hydrolysis is a long‐standing challenge in catalysis. For prophylactic treatment against organophosphate exposure, enzymatic hydrolysis needs to occur at high rates in the presence of low substrate concentrations and enzymatic activity should persist over days and weeks. Here, the conjugation of small DNA scaffolds was used to introduce substrate binding sites with micromolar affinity to VX, paraoxon, and methyl‐parathion in close proximity to the enzyme phosphotriesterase (PTE). The result was a decrease in KM and increase in the rate at low substrate concentrations. An optimized system for paraoxon hydrolysis decreased KM by 11‐fold, with a corresponding increase in second‐order rate constant. The initial rates of VX and methyl‐parathion hydrolysis were also increased by 3.1‐ and 6.7‐fold, respectively. The designed scaffolds not only increased the local substrate concentration, but they also resulted in increased stability and PTE‐DNA particle size tuning between 25 and ~150 nm. The scaffold engineering approach taken here is focused on altering the local chemical and physical microenvironment around the enzyme and is therefore compatible with active site engineering via combinatorial and computational approaches.

Local Environment Affects the Activity of Enzymes on a 3D Molecular Scaffold

“…In order to explain the enhancement of the catalytic kinetics of a single enzyme, the pH profile near the DNA surface was modeled, and it was concluded that the pH might have a critical effect on enzymatic activity . Moreover, research also showed that HRP − and phosphotriesterase’s , activities can be improved by increasing the affinity of substrates on negatively charged DNA motifs, implying that the polyanionic DNA material could promote enzymatic activity through altering local chemical and physical microenvironments.…”

Multienzymatic Cascades and Nanomaterial Scaffolding—A Potential Way Forward for the Efficient Biosynthesis of Novel Chemical Products

“…[47][48][49][50] However, here the underlying contributing factors are more complex and appear to be associated with the influence of DNA's strongly charged environment leading to localized sequestration of substrate or intermediaries along with perhaps making the reaction environment more favorable in certain cases. [51,52] There has been, and still continues to be, considerable scientific debate around the origins and complexity surrounding such phenomena. [53,54] Previous studies of enzymatic enhancement on NP scaffolds in the context of a coupled enzymatic system revealed that enzyme stabilization could also contribute to the observed catalytic increases.…”

Enzymes, Reacting with Organophosphorus Compounds as Detoxifiers: Diversity and Functions

“…with N-acyl homoserine lactones [108,109]. Thus, they can be rationally combined to obtain universal enzyme preparations [110] or even hybrid molecules with other enzymes [111] or with 'guide' DNA [112]. This opens up new promising perspectives for their practical application.…”