Mutation of residues in the coenzyme binding pocket of Dopa decarboxylase. Effects on catalytic properties.

摘要: Residues D271, H192, H302 and N300 of l-3,4-dihydroxyphenylalanine decarboxylase (DDC), a homodimeric pyridoxal 5′-phosphate (PLP) enzyme, were mutated in order to acquire information on the catalytic mechanism. These residues are potential participants catalysis because they belong common PLP-binding structural motif group I, II III decarboxylases other PLP enzymes, among putative active-site modelled rat liver DDC. The spectroscopic features D271E, H192Q, H302Q N300A mutants as well as their dissociation constants for suggest that substitution each these causes alteration state bound coenzyme molecule and of conformation aromatic amino acids, possibly in vicinity active site. This supports, but does not prove, possibility located coenzyme-binding cleft. Interestingly, mutation residue generates an oxidative activity towards (l-Dopa), inherent wild-type aerobiosis, reduces nonoxidative l-Dopa from 3- 390-fold. partition ratio between decarboxylation ranges 5.7 × 10−4 mutant 946 × 10−4 mutant. Unlike mutants catalyse two reactions same extent either presence or absence O2. In addition, all four exhibit extremely low level deaminase serotonin with respect wild-type. All findings demonstrate although essential catalysis, alters nature catalysis. A possible relationship integrity cleft, productive binding O2 transition closed conformational DDC is discussed.