Cloning and expression of Escherichia coli 5'-methylthioadenosine/S-adenosylhomocysteine nucleosidase: identification of the pfs gene product

Abstract

The enzyme 5'-methylthioadenosine/S-adenosylhomocysteine nucleosidase (EC 3.2.2.9) is responsible for cleavage of the glycosidic bond in both 5'-methylthioadenosine (MTA) and S-adenosylhomocysteine (SAH). Based on amino acid sequence analysis of this enzyme from Klebsiella, we recently speculated that an open reading frame found in E. coli (designated pfs) encoded MTA/SAH nucleosidase. To explore this possibility, we amplified, cloned, and expressed the complete pfs gene from E. coli genomic DNA. The recombinant protein exhibited a molecular weight and Michaelis constants for MTA that are in agreement with those reported for native enzyme. From this biochemical evidence we confirm our original assignment of the pfs gene as encoding MTA/SAH nucleosidase.