Mechanism and Catalytic Site Atlas

Introduction

The reaction comprises of two steps, both involving nucleophilic attack. The first step starts by the addition of the carbon atom of the guanidino group of L-Arg to the thiol group of Cys407 of the enzyme, and donation of the thiol proton to the L-Arg substrate. The thiol group of C407 forms a covalently bound intermediate with the carbon atom of the guanidino group, and donates its proton to the imino group of the arginine substrate. Hydrogen bonds and salt linkage of the guanidino nitrogens to Asp170 and Asp305 enhance the electrophilicity of the guanidino-carbon atom, and a potential hydrogen bond of the epsilon-imino nitrogen to His303 supports proton transfer to this atom. Subsequently the bond between the epsilon-imino group and the amidino-carbon atom is broken to generate L-Orn. The second step starts with a proton transfer from the positively charged Gly substrate to His303 and a nucleophilic attack of the lone electron pair of the Gly nitrogen atom at the carbon atom of the bound amidino group. This is followed by the formation of a tetrahedral adduct and its collapse by cleavage of the amidino-carbon-sulfur bond to generate guanidinoacetic acid.

The amidino-carbon atom of the arginine substrate is placed between the thiol group of the cysteine residue and the imino group of the histidine residue. The arginine is fixed tightly to the active site by formation of hydrogen bonds between the amidino nitrogen atoms and the carboxyl groups of aspartate residues 170 and 305. These orient the substrate's guanidino group such that the sulphur atom of CYS 407, the guanidino-carbon atom and the nitrogen atom of HIS 303 are in a line orthogonal to the plane of the guanidino group.