Elucidation of mechanisms for catalytic center maturation and catalysis of nitrile hydratase family enzyme

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By performing time-resolved X-ray crystallography of Fe-type nitrile hydratase(NHase) whoseβArg56 were substituted by Lys, we obtained the structure of the enzyme-substrate complex. By replacing two Arg residues resided in the active site pocket of a NHase family enzyme, thiocyanate hydrolase(SCNase), we showed that the size as well as the surface charge of the substrate binding pocket was important for the substrate specificities of SCNase. As for the maturation mechanism, we found that the SCNase activator protein, P15K, is responsible for the refolding, metal-insertion and post-translational modification of Cys ligand in the catalytic subunit,γ. The metal-incorporatedγsubunit form the complex with the other subunits,αandβ, to form the mature SCNase.