Enhancing Paraoxon Binding to Organophosphorus Hydrolase Active Site

El Khoury, Léa; Mobley, David L.; Ye, Dongmei; Rempe, Susan B.

doi:10.3390/ijms222312624

Enhancing Paraoxon Binding to Organophosphorus Hydrolase Active Site

Journal Article · Mon Nov 22 23:00:00 EST 2021 · International Journal of Molecular Sciences (Online)

DOI:https://doi.org/10.3390/ijms222312624· OSTI ID:1833102

; ; Ye, Dongmei; Rempe, Susan B.

Organophosphorus hydrolase (OPH) is a metalloenzyme that can hydrolyze organophosphorus agents resulting in products that are generally of reduced toxicity. The best OPH substrate found to date is diethyl p-nitrophenyl phosphate (paraoxon). Most structural and kinetic studies assume that the binding orientation of paraoxon is identical to that of diethyl 4-methylbenzylphosphonate, which is the only substrate analog co-crystallized with OPH. In the current work, we used a combined docking and molecular dynamics (MD) approach to predict the likely binding mode of paraoxon. Then, we used the predicted binding mode to run MD simulations on the wild type (WT) OPH complexed with paraoxon, and OPH mutants complexed with paraoxon. Additionally, we identified three hot-spot residues (D253, H254, and I255) involved in the stability of the OPH active site. We then experimentally assayed single and double mutants involving these residues for paraoxon binding affinity. The binding free energy calculations and the experimental kinetics of the reactions between each OPH mutant and paraoxon show that mutated forms D253E, D253E-H254R, and D253E-I255G exhibit enhanced substrate binding affinity over WT OPH. Interestingly, our experimental results show that the substrate binding affinity of the double mutant D253E-H254R increased by 19-fold compared to WT OPH.

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Sponsoring Organization:: USDOE

OSTI ID:: 1833102

Alternate ID(s):: OSTI ID: 1838185

Journal Information:: International Journal of Molecular Sciences (Online), Journal Name: International Journal of Molecular Sciences (Online) Journal Issue: 23 Vol. 22; ISSN 1422-0067; ISSN IJMCFK

Publisher:: MDPI AGCopyright Statement

Country of Publication:: Switzerland

Language:: English

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