Mycobacteria express enzymes from both the de novo and purine-salvage pathways. However, the regulation of these processes and the roles of individual metabolic enzymes have not been sufficiently detailed. Both Mycobacterium tuberculosis (Mtb) and Mycobacterium smegmatis (Msm) possess three guaB genes, but information is only available on guaB2, which encodes an essential inosine 5'-monophosphate dehydrogenase (IMPDH) involved in de novo purine biosynthesis. This study shows that guaB1, annotated in databases as a putative IMPDH, encodes a guanosine 5'-monophosphate reductase (GMPR), which recycles guanosine monophosphate to inosine monophosphate within the purine-salvage pathway and contains a cystathionine-β-synthase domain (CBS), which is essential for enzyme activity. GMPR activity is allosterically regulated by the ATP/GTP ratio in a pH-dependent manner. Bioinformatic analysis has indicated the presence of GMPRs containing CBS domains across the entire Actinobacteria phylum.

• Keywords
• CBS domain, GMPR, Mycobacterium, guaB1, purine biosynthesis,
• MeSH
• Adenosine Triphosphate MeSH
• Cystathionine beta-Synthase genetics   MeSH
• Cystathionine * MeSH
• GMP Reductase genetics   metabolism   MeSH
• Guanosine Triphosphate MeSH
• IMP Dehydrogenase genetics   metabolism   MeSH
• Inosine MeSH
• Inosine Monophosphate metabolism   MeSH
• Guanosine Monophosphate metabolism   MeSH
• Mycobacterium tuberculosis * metabolism   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Adenosine Triphosphate MeSH
• Cystathionine beta-Synthase MeSH
• Cystathionine * MeSH
• GMP Reductase MeSH
• Guanosine Triphosphate MeSH
• IMP Dehydrogenase MeSH
• Inosine MeSH
• Inosine Monophosphate MeSH
• Guanosine Monophosphate MeSH

Purine metabolism plays a ubiquitous role in the physiology of Mycobacterium tuberculosis and other mycobacteria. The purine salvage enzyme hypoxanthine-guanine phosphoribosyltransferase (HGPRT) is essential for M. tuberculosis growth in vitro; however, its precise role in M. tuberculosis physiology is unclear. Membrane-permeable prodrugs of specifically designed HGPRT inhibitors arrest the growth of M. tuberculosis and represent potential new antituberculosis compounds. Here, we investigated the purine salvage pathway in the model organism Mycobacterium smegmatis Using genomic deletion analysis, we confirmed that HGPRT is the only guanine and hypoxanthine salvage enzyme in M. smegmatis but is not required for in vitro growth of this mycobacterium or survival under long-term stationary-phase conditions. We also found that prodrugs of M. tuberculosis HGPRT inhibitors displayed an unexpected antimicrobial activity against M. smegmatis that is independent of HGPRT. Our data point to a different mode of mechanism of action for these inhibitors than was originally proposed.IMPORTANCE Purine bases, released by the hydrolytic and phosphorolytic degradation of nucleic acids and nucleotides, can be salvaged and recycled. The hypoxanthine-guanine phosphoribosyltransferase (HGPRT), which catalyzes the formation of guanosine-5'-monophosphate from guanine and inosine-5'-monophosphate from hypoxanthine, represents a potential target for specific inhibitor development. Deletion of the HGPRT gene (Δhgprt) in the model organism Mycobacterium smegmatis confirmed that this enzyme is not essential for M. smegmatis growth. Prodrugs of acyclic nucleoside phosphonates (ANPs), originally designed against HGPRT from Mycobacterium tuberculosis, displayed anti-M. smegmatis activities comparable to those obtained for M. tuberculosis but also inhibited the ΔhgprtM. smegmatis strain. These results confirmed that ANPs act in M. smegmatis by a mechanism independent of HGPRT.

• Keywords
• Mycobacterium smegmatis, guanine, hypoxanthine, hypoxanthine-guanine phosphoribosyltransferase, inhibitors, purine salvage pathway,
• MeSH
• Antitubercular Agents chemistry   pharmacology   MeSH
• Hypoxanthine Phosphoribosyltransferase antagonists & inhibitors   chemistry   genetics   metabolism   MeSH
• Enzyme Inhibitors chemistry   pharmacology   MeSH
• Catalysis MeSH
• Metabolic Networks and Pathways MeSH
• Microbial Viability MeSH
• Mycobacterium smegmatis genetics   growth & development   metabolism   MeSH
• Plasmids genetics   MeSH
• Purines metabolism   MeSH
• Dose-Response Relationship, Drug MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Antitubercular Agents MeSH
• Hypoxanthine Phosphoribosyltransferase MeSH
• Enzyme Inhibitors MeSH
• purine MeSH Browser
• Purines MeSH