Adduct of the plant protein RuBisCO formed upon interaction with G-type organophosphorus nerve agents

Abstract

The Chemical Weapons Prohibition Convention (CWC) explicitly prohibits the use of organophosphorus nerve agents (OPNAs), which are highly toxic cholinesterase inhibitors that pose a serious threat to human peace, health and security. Currently, protein adducts of OPNAs are mainly studied in humans and animals. However, a novel and reliable biomarker must be found immediately to provide retrospective identification of OPNA exposure in environmental samples. In this study, we aimed to broaden the detection window for nerve agents in various species, explore prospective biomarkers for their high reactivity and good stability, and examine the feasibility of adding plant proteins to G-type OPNAs. We investigated the plant protein ribulose-1,5-diphosphate carboxylated oxygenase (RuBisCO). Several modification sites were discovered, including S115/S117 and Y118/Y120 on EHHNS*PGY*Y*DGR and K175 on Y*GRPLLGCTIK*PK. Ten reliable and stable peptide segments were also identified. Both tabun (GA) and soman (GD) can concurrently modify the peptides EHHNS*PGY*Y*DGR, WS*PELAAACEVWK* , Y*GRPLLGCTIK*PK, and GHYLNATAGTCEDMMK. As a result, the exposure to OPNAs can be retrospectively verified using these stabilized peptides. The stability of the three OPNA adducts varies. The sarin (GB) adduct is the most stable, while GA-Lys and GA-Ser show the fastest aging rate, followed by GD. The differential aging rates of OPNA adducts may provide temporal clues for distinguishing exposure sources, provided that aging kinetics are validated in vivo and under specified post-exposure timeframes. This study proposes a novel method for retrospective organophosphorus analysis, with potential applications in forensic investigations and chemical weapon verification. A computer-based molecular simulation of the highly reactive stable modification sites revealed that hydrogen bonds are the driving force for the formation of covalent compounds. This study contributes to our understanding of the mechanism of modification site formation of the RuBisCO adduct.