Role of zinc ion for catalytic activity in d -serine dehydratase from Saccharomyces cerevisiae

Abstract

d-Serine dehydratase from Saccharomyces cerevisiae (DsdSC) is a fold-type III pyridoxal 5′-phosphate-dependent enzyme catalyzing d-serine dehydration. The enzyme contains 1 mol Zn 2+ in its active site and shows a unique zinc dependence. The Zn 2+ is essential for the d-serine dehydration, but not for the α,β-elimination of β-Cl-d-alanine catalyzed as a side-reaction. The fact that dehydration of d-threonine and d-allo-threonine, also catalyzed by DsdSC, is likewise Zn 2+ dependent indicates that Zn 2+ is indispensable for the elimination of hydroxyl group, regardless of the stereochemistry of C β. Removal of Zn 2+ results in a less polar active site without changing the gross conformation of DsdSC. 1H NMR determined the rates of α-hydrogen abstraction and hydroxyl group elimination of d-serine in 2H 2O to be 9.7 and 8.5 s -1, respectively, while the removal of Zn 2+ abolished both reactions. Mutation of Cys400 or His398 within the Zn 2+ binding sites to Ala endowed DsdSC with similar properties to those of the Zn 2+-depleted wild-type enzyme: the mutants lost the reactivity toward d-serine and d-threonine but retained that toward β-Cl-d-alanine. 1H NMR analysis also revealed that both α-hydrogen abstraction and hydroxyl group elimination from d-serine were severely hampered in the C400A mutant. Our data suggest that DsdSC catalyzes the α-hydrogen abstraction and hydroxyl group elimination in a concerted fashion. © 2011 The Authors Journal compilation © 2011 FEBS.