Direct Sulfhydrylation for Methionine Biosynthesis in Leptospira meyeri

A gene library of theLeptospira meyeri serovar semaranga strain Veldrat S.173 DNA has been constructed in a mobilizable cosmid with inserts of up to 40 kb. It was demonstrated that aLeptospira DNA fragment carryingmetY complementedEscherichia coli strains carrying mutations inmetB . The latter gene encodes cystathionine γ-synthase, an enzyme which catalyzes the second step of the methionine biosynthetic pathway. ThemetY gene is 1,304 bp long and encodes a 443-amino-acid protein with a molecular mass of 45 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The deduced amino acid sequence of theLeptospira metY product has a high degree of similarity to those ofO -acetylhomoserine sulfhydrylases fromAspergillus nidulans andSaccharomyces cerevisiae . A lower degree of sequence similarity was also found with bacterial cystathionine γ-synthase. TheL. meyeri metY gene was overexpressed under the control of the T7 promoter. MetY exhibits anO -acetylhomoserine sulfhydrylase activity. Genetic, enzymatic, and physiological studies reveal that the transsulfuration pathway via cystathionine does not exist inL. meyeri , in contrast to the situation found for fungi and some bacteria. Our results indicate, therefore, that theL. meyeri MetY enzyme is able to perform direct sulfhydrylation for methionine biosynthesis by usingO -acetylhomoserine as a substrate.