Theoretcial Investigation on Reaction Mechanism of Binuclear Nickel Guanidine Hydrolase

Abstract The nitrogen‐rich compound guanidine is widely distributed in nature, but its utilization is hindered by strong resonance stabilization. GdmH, a binuclear nickel enzyme from Synechocystis sp. PCC 6803, is capable of directly converting guanidinium cation into urea and ammonium. In this study, we employed density functional calculations to investigate the reaction mechanism of GdmH using a chemical model derived from the enzyme's X‐ray crystal structure. The calculations revealed that the GdmH‐catalyzed guanidinium hydrolysis proceeds through a nucleophilic attack by the di‐nickel bridging hydroxide on the guanidinium carbon forming a tetrahedral intermediate, two proton transfer steps from the hydroxyl to an amino facilitated by Asp203, C─N bond cleavage yielding urea and ammonia, and regeneration of the bridging hydroxide accompanied by ammonium release. The rate‐limiting step is the first proton transfer from hydroxyl to Asp203, with an energy barrier of 11.8 kcal mol −1 . Comparative analyses demonstrated that neutral guanidine cannot be hydrolyzed by GdmH due to the absence of a positive charge, which is essential for effective catalysis. Further investigations showed that GdmH is inefficient in catalyzing urea hydrolysis. These findings enhance our understanding of the catalytic specificity of GdmH and the role of nickel cofactors in biological enzymatic processes.