Nucleoside diphosphate kinase-like activity in adenylate kinase of Mycobacterium tuberculosis

Abstract

Ak (adenylate kinase) is a ubiquitous enzyme that catalyses a reversible high-energy phosphoryl-transfer reaction between ATP and AMP to form ADP. In the present study, the Ak gene (adk) of mycobacterium tuberculosis was cloned, expressed in escherichia coli and purified as a glutathione S-transferase fusion protein. Purified Ak converted AMP into ADP in the presence of [γ -³²P]ATP or [γ -³²P]GTP. Replacement of arginine-88 of adk with glycine resulted in the loss of enzymic activity. The purified protein also showed Ndk (nucleoside diphosphate kinase)-like activity as it transferred terminal phosphate from [γ-³²P]ATP to all nucleoside diphosphates, converting them into corresponding triphosphates. However, Ndk-like activity of Ak was not observed with [γ-³²P]GTP. Immunoblot analysis of various cellular fractions of M. tuberculosis H₃₇Rv revealed that Ak is a cytoplasmic protein. The dual activity of Ak as both nucleoside mono- and di-phosphate kinases suggested that this enzyme may have a role in RNA and DNA biosynthesis in addition to its role in intracellular nucleotide metabolism.