Theoretical Studies On The Reaction Mechanism Of Protein S-Nitrosylation

Protein S-nitrosylation, the covalent binding of nitric oxide (NO·) to the thiol group of cysteine, has emerged as a redox-based post-translational modification and conveyed the ubiquitous influence of nitric oxide on cellular signal transduction. Increasing evidences support that S-nitrosylation is related to many diseases. In the early literature, S-nitrosylation is generally found to produce S-nitrosothiol (RSNO). Recently, it is also reported that S-nitroxide (RS-NH-O·) may be another stable form in S-nitrosylated hemoglobin (Hb). Here we have conducted quantum mechanical (QM) molecular dynamics simulations CPMD, with lowest free energy reaction pathway search method"metadynamics", to investigate the chemical behaviors of RSH, RS-NH-O·and RSNO in aqueous solution and methanol, respectively. Based on the computational results of CPMD-Metadynamics, we propose a purely NO·-dependent S-nitrosylation reaction mechanism, in which⑴deprotonation of thiol is the rate determining step;⑵thiol anion RS- and radical RS·capture NO·at different stages; and⑶no additional electron acceptor is needed during the process. In addition, RS-NH-O·is verified as an intermediate of S-nitrosylation. In less protolytic surroundings, its S-N bond will experience homolytic cleavage, producing nitroxyl (HNO) and RS·radical. RS·radical can combine another molecule of NO·, leading to the final product RSNO. Overall, in our calculations, the S-nitrosylation occurs in terms of RSH + 2 NO·RSNO + HNO.To further investigate the importance of thiol deprotonation in protein S-nitrosylation, molecular mechanical (MM) molecular dynamics simulations have been employed to study the local environments of the modified sites in R/T state of Hb. Compared to T state, R state is considered to be modified more feasibly. The MM simulations show that Hisβ92 and Tyrβ145 might involve in Cysβ93 thiol deprotonation.We also carried out Structural Bioinformatics studies on the proteins which have been reported to be S-nitrosylated. The statistical analysis shows that S-nitrosylated cysteines are in hydrophilic regions when compared to negative collection.