Molecular binding scaffolds increase local substrate concentration enhancing the enzymatic hydrolysis of VX nerve agent.,Biotechnology and Bioengineering

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Molecular binding scaffolds increase local substrate concentration enhancing the enzymatic hydrolysis of VX nerve agent.
Biotechnology and Bioengineering ( IF 3.5 ) Pub Date : 2020-04-01 , DOI: 10.1002/bit.27346
Xuye Lang ₁ , Xiao Hong ₂ , Cetara A Baker ₃ , Tamara C Otto ₃ , Ian Wheeldon _{1,

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Affiliation

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 K _M and increase in the rate at low substrate concentrations. An optimized system for paraoxon hydrolysis decreased K _M 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.

中文翻译：

分子结合支架增加了局部底物浓度，增强了 VX 神经毒剂的酶水解。

有机磷酸酯水解的动力学增强是催化领域的一个长期挑战。对于有机磷暴露的预防性治疗，酶促水解需要在低底物浓度下以高速率发生，酶促活性应持续数天和数周。在这里，小 DNA 支架的结合用于在酶磷酸三酯酶 (PTE) 附近引入对 VX、对氧磷和甲基对硫磷具有微摩尔亲和力的底物结合位点。结果是在低底物浓度下K _M降低和速率增加。对氧磷水解的优化系统降低了K _M11 倍，二阶速率常数相应增加。VX 和甲基对硫磷水解的初始速率也分别增加了 3.1 倍和 6.7 倍。设计的支架不仅增加了局部底物浓度，而且还提高了稳定性和 PTE-DNA 粒径在 25 和 ~150 nm 之间的调整。这里采用的支架工程方法侧重于改变酶周围的局部化学和物理微环境，因此通过组合和计算方法与活性位点工程兼容。

更新日期：2020-04-01

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