Hypoxanthine-Guanine Phosphoribosyltransferase Is Dispensable for Mycobacterium smegmatis Viability

Knejzlı́k, Zdeněk; Herkommerová, Klára; Hocková, Dana; Pichová, Iva

doi:10.1128/jb.00710-19

Cited by 6 publications

(8 citation statements)

References 30 publications

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“…Next, we blocked the de novo purine metabolic pathway by knocking out the essential purF gene, which encodes amidophosphoribosyltransferase, catalysing the first step of purine de novo biosynthesis (Fig. 1A ), and compared the growth of the ΔguaB1ΔpurF strain with wt and ΔpurF Msm strains [ 43 , 44 ]. For its growth, the ΔpurF strain needed an external source of purine such as hypoxanthine, adenine or guanine (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…2B ), which serve as precursors for purine metabolism via the corresponding salvaging enzymes (Fig. 1A ) [ 44 ]. The ΔguaB1ΔpurF strain grew on media containing adenine or hypoxanthine but not on media containing guanine as the sole purine source (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The pFLAG‐Msm.GuaB1 (L‐ΔCBS) encoding GuaB1 lacking the region V96‐L194, with it being replaced by the SGG motif, was constructed as follows: pFLAG‐Msm.GuaB1 was PCR linearised by the primer pair 15/16 and circularised by the in‐fusion approach. The plasmid encoding of C‐terminally His‐tagged Mtb GMPR for expression in Msm was based on the pSE200 vector [ 44 ], derived from the pSE100 plasmid containing a constitutive Pmyc1tetO ‐promoter in the absence of a Tet repressor [ 75 ]. Mtb.guaB1His was amplified from genomic DNA by PCR using the primer pair 21/22 and inserted into Swa I‐linearised pSE200 by the in‐fusion approach.…”

Section: Methodsmentioning

confidence: 99%

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The mycobacterial guaB1 gene encodes a guanosine 5′‐monophosphate reductase with a cystathionine‐β‐synthase domain

et al. 2022

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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.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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The mycobacterial guaB1 gene encodes a guanosine 5′‐monophosphate reductase with a cystathionine‐β‐synthase domain

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“…The purine salvage enzyme HGPRT was found essential for Mycobacterium tuberculosis growth in vitro ; however, its precise role in M. tuberculosis physiology is so far unclear. Membrane permeable prodrugs of HGPRT inhibitors arrest the growth of M. tuberculosis and represent potential new antituberculosis compounds ( Knejzlík et al, 2020 ).…”

Section: Acyclic Nucleoside Phosphonates (Anps)mentioning

confidence: 99%

Phosphonates and Phosphonate Prodrugs in Medicinal Chemistry: Past Successes and Future Prospects

et al. 2022

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Compounds with a phosphonate group, i.e., –P(O)(OH)2 group attached directly to the molecule via a P-C bond serve as suitable non-hydrolyzable phosphate mimics in various biomedical applications. In principle, they often inhibit enzymes utilizing various phosphates as substrates. In this review we focus mainly on biologically active phosphonates that originated from our institute (Institute of Organic Chemistry and Biochemistry in Prague); i.e., acyclic nucleoside phosphonates (ANPs, e.g., adefovir, tenofovir, and cidofovir) and derivatives of non-nucleoside phosphonates such as 2-(phosphonomethyl) pentanedioic acid (2-PMPA). Principal strategies of their syntheses and modifications to prodrugs is reported. Besides clinically used ANP antivirals, a special attention is paid to new biologically active molecules with respect to emerging infections and arising resistance of many pathogens against standard treatments. These new structures include 2,4-diamino-6-[2-(phosphonomethoxy)ethoxy]pyrimidines or so-called “open-ring” derivatives, acyclic nucleoside phosphonates with 5-azacytosine as a base moiety, side-chain fluorinated ANPs, aza/deazapurine ANPs. When transformed into an appropriate prodrug by derivatizing their charged functionalities, all these compounds show promising potential to become drug candidates for the treatment of viral infections. ANP prodrugs with suitable pharmacokinetics include amino acid phosphoramidates, pivaloyloxymethyl (POM) and isopropoxycarbonyloxymethyl (POC) esters, alkyl and alkoxyalkyl esters, salicylic esters, (methyl-2-oxo-1,3-dioxol-4-yl) methyl (ODOL) esters and peptidomimetic prodrugs. We also focus on the story of cytostatics related to 9-[2-(phosphonomethoxy)ethyl]guanine and its prodrugs which eventually led to development of the veterinary drug rabacfosadine. Various new ANP structures are also currently investigated as antiparasitics, especially antimalarial agents e.g., guanine and hypoxanthine derivatives with 2-(phosphonoethoxy)ethyl moiety, their thia-analogues and N-branched derivatives. In addition to ANPs and their analogs, we also describe prodrugs of 2-(phosphonomethyl)pentanedioic acid (2-PMPA), a potent inhibitor of the enzyme glutamate carboxypeptidase II (GCPII), also known as prostate-specific membrane antigen (PSMA). Glutamate carboxypeptidase II inhibitors, including 2-PMPA have been found efficacious in various preclinical models of neurological disorders which are caused by glutamatergic excitotoxicity. Unfortunately its highly polar character and hence low bioavailability severely limits its potential for clinical use. To overcome this problem, various prodrug strategies have been used to mask carboxylates and/or phosphonate functionalities with pivaloyloxymethyl, POC, ODOL and alkyl esters. Chemistry and biological characterization led to identification of prodrugs with 44–80 fold greater oral bioavailability (tetra-ODOL-2-PMPA).

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“…Next, we blocked the de novo purine metabolic pathway by knocking out the essential purF gene and compared the growth of the guaB1purF strain with that of the wt and purF Msm strains (Knejzlik et al, 2020;Knejzlik et al, 2019). The purF strain requires for growth an external source of purine such as hypoxanthine, adenine or ,guanine to serve as a precursor for purine metabolism via the corresponding salvaging enzymes (Knejzlik et al, 2020). The guaB1purF strain grew on media containing adenine or hypoxanthine but not on media containing guanine as the sole purine source (Fig.…”

Section: Msm Guab1 Is An Nadph-dependent Gmpr Involved In the Purine Salvage Pathwaymentioning

confidence: 99%

The mycobacterial guaB1 gene encodes a guanosine 5’-monophosphate reductase with a cystathione-β-synthase domain

Knejzlı́k

Doležal

Herkommerová

et al. 2021

Preprint

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Purine metabolism plays a pivotal role in bacterial life cycle, however, regulation of the de novo and purine salvage pathways have not been extensively detailed in mycobacteria. By gene knockout, biochemical and structural analyses, we identified Mycobacterium smegmatis (Msm) and Mycobacterium tuberculosis (Mtb) guaB1 gene product as a novel type of guanosine 5’-monophosphate reductase (GMPR), which recycles guanosine monophosphate to inosine monophosphate within the purine salvage pathway and contains cystathione β-synthase (CBS) domains with atypical orientation in the octamer. CBS domains share a much larger interacting area with a conserved catalytic domain in comparison with the only known CBS containing protozoan GMPR and closely related inosine monophosphate dehydrogenase structures. Our results revealed essential effect of pH on allosteric regulation of Msm GMPR activity and oligomerization with adenine and guanosine nucleotides binding to CBS domains.Bioinformatic analysis indicated the presence of GMPRs containing CBS domains across the entire Actinobacteria phylum.

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Hypoxanthine-Guanine Phosphoribosyltransferase Is Dispensable for Mycobacterium smegmatis Viability

Cited by 6 publications

References 30 publications

The mycobacterial guaB1 gene encodes a guanosine 5′‐monophosphate reductase with a cystathionine‐β‐synthase domain

The mycobacterial guaB1 gene encodes a guanosine 5′‐monophosphate reductase with a cystathionine‐β‐synthase domain

Phosphonates and Phosphonate Prodrugs in Medicinal Chemistry: Past Successes and Future Prospects

The mycobacterial guaB1 gene encodes a guanosine 5’-monophosphate reductase with a cystathione-β-synthase domain

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