Kinetic Mechanism of Nicotinic Acid Phosphoribosyltransferase:  Implications for Energy Coupling

Gross, Janet L.; Rajavel, Mathumathi; Grubmeyer, Charles

doi:10.1021/bi972014w

Cited by 34 publications

(37 citation statements)

References 24 publications

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“…The same strict substrate preference for Nm over Na (ϳ5,000-fold) was observed by us for the recombinant purified abNadV enzyme (Table 3), confirming its role in the nondeamidating route. Notably, both phosphoribosyltransferases, abPncB and abNadV, are strongly activated in the presence of 2 mM ATP (ϳ100-and ϳ10-fold, respectively) as previously reported for other representatives of these families (43,44). Comparison of catalytic efficiencies of abNadV and abPncA (Table 3) potentially competing for the Nm substrate suggests that the deamidating route is likely to prevail, especially at low concentrations of Nm (as the K m value of abNadV is ϳ 20-fold higher than of abPncA).…”

Section: Table 2 Growth Phenotypes Of a Baylyi Adp1 Knockout Mutantssupporting

confidence: 80%

Genomics-driven Reconstruction of Acinetobacter NAD Metabolism

Sorci

Blaby

Ingeniis

et al. 2010

Journal of Biological Chemistry

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Enzymes involved in the last steps of NAD biogenesis, nicotinate mononucleotide adenylyltransferase (NadD) and NAD synthetase (NadE), are conserved and essential in most bacterial species and are established targets for antibacterial drug development. Our genomics-based reconstruction of NAD metabolism in the emerging pathogen Acinetobacter baumannii revealed unique features suggesting an alternative targeting strategy. Indeed, genomes of all analyzed Acinetobacter species do not encode NadD, which is functionally replaced by its distant homolog NadM. We combined bioinformatics with genetic and biochemical techniques to elucidate this and other important features of Acinetobacter NAD metabolism using a model (nonpathogenic) strain Acinetobacter baylyi sp. ADP1. Thus, a comparative kinetic characterization of PncA, PncB, and NadV enzymes allowed us to suggest distinct physiological roles for the two alternative, deamidating and nondeamidating, routes of nicotinamide salvage/recycling. The role of the NiaP transporter in both nicotinate and nicotinamide salvage was confirmed. The nondeamidating route was shown to be transcriptionally regulated by an ADP-ribose-responsive repressor NrtR. The NadM enzyme was shown to possess dual substrate specificity toward both nicotinate and nicotinamide mononucleotide substrates, which is consistent with its essential role in all three routes of NAD biogenesis, de novo synthesis as well as the two salvage pathways. The experimentally confirmed unconditional essentiality of nadM provided support for the choice of the respective enzyme as a drug target. In contrast, nadE, encoding a glutamine-dependent NAD synthetase, proved to be dispensable when the nondeamidating salvage pathway functioned as the only route of NAD biogenesis.Acinetobacter baumannii is an emerging pathogen that belongs to a relatively underexplored branch of ␥-proteobacteria. It can cause severe pneumonia and infections of the urinary tract, bloodstream, and other parts of the body. Some isolates of A. baumannii display resistance to many known antibiotics (1, 2), emphasizing the importance of pursuing new therapeutic targets for drug development. Biogenesis of nicotinamide adenine nucleotide (NAD), an indispensable cofactor involved in a multitude of biochemical transformations in metabolic networks of all species, was recently established as a target pathway for the development of new antibiotics (3-7). Beyond its main function as a redox cofactor, NAD is consumed as a co-substrate by a number of nonredox enzymes such as bacterial DNA ligase and protein deacetylase of the CobB/Sir2 family (8, 9). A degradative consumption of NAD by these and, likely, other (not fully elucidated) enzymes demands continuous replenishing of the NAD pool, providing further rationale for targeting essential enzymes involved in its biogenesis and recycling. Among these enzymes, nicotinate mononucleotide adenylyltransferase (NaMNAT) 4 of the NadD family and NAD synthetase of the NadE family are widely recognized as the most promising...

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Section: Table 2 Growth Phenotypes Of a Baylyi Adp1 Knockout Mutantssupporting

confidence: 80%

Genomics-driven Reconstruction of Acinetobacter NAD Metabolism

Sorci

Blaby

Ingeniis

et al. 2010

Journal of Biological Chemistry

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“…That is, the enzyme is phosphorylated by ATP at His219, which results in the formation of a high-affinity form with K m values for nicotinate and PRPP 200-fold lower than those of the unphosphorylated enzyme. Also, the coupling of ATP hydrolysis to the phosphoribosyltransferase reaction causes a change in the equilibrium value, from 0.67 to 1,100 (253)(254)(255). Mutational replacement of His219 completely abrogates phosphorylation and stimulation by ATP (256).…”

Section: Reactions At the Anomeric Carbon Of Prppmentioning

confidence: 99%

Phosphoribosyl Diphosphate (PRPP): Biosynthesis, Enzymology, Utilization, and Metabolic Significance

Hove‐Jensen

Andersen

Kilstrup

et al. 2017

Microbiol Mol Biol Rev

168

187

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SUMMARY Phosphoribosyl diphosphate (PRPP) is an important intermediate in cellular metabolism. PRPP is synthesized by PRPP synthase, as follows: ribose 5-phosphate + ATP → PRPP + AMP. PRPP is ubiquitously found in living organisms and is used in substitution reactions with the formation of glycosidic bonds. PRPP is utilized in the biosynthesis of purine and pyrimidine nucleotides, the amino acids histidine and tryptophan, the cofactors NAD and tetrahydromethanopterin, arabinosyl monophosphodecaprenol, and certain aminoglycoside antibiotics. The participation of PRPP in each of these metabolic pathways is reviewed. Central to the metabolism of PRPP is PRPP synthase, which has been studied from all kingdoms of life by classical mechanistic procedures. The results of these analyses are unified with recent progress in molecular enzymology and the elucidation of the three-dimensional structures of PRPP synthases from eubacteria, archaea, and humans. The structures and mechanisms of catalysis of the five diphosphoryltransferases are compared, as are those of selected enzymes of diphosphoryl transfer, phosphoryl transfer, and nucleotidyl transfer reactions. PRPP is used as a substrate by a large number phosphoribosyltransferases. The protein structures and reaction mechanisms of these phosphoribosyltransferases vary and demonstrate the versatility of PRPP as an intermediate in cellular physiology. PRPP synthases appear to have originated from a phosphoribosyltransferase during evolution, as demonstrated by phylogenetic analysis. PRPP, furthermore, is an effector molecule of purine and pyrimidine nucleotide biosynthesis, either by binding to PurR or PyrR regulatory proteins or as an allosteric activator of carbamoylphosphate synthetase. Genetic analyses have disclosed a number of mutants altered in the PRPP synthase-specifying genes in humans as well as bacterial species.

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“…Although not required for the synthesis of NMN from NAm and 5-phospho-D-ribosyl-1-pyrophosphate (PRPP), energy from the ATPase activity improves catalysis and lowers the K m for NAm. Previously demonstrated only for human and bacterial (16,26) enzymes, phage NAmPRTases such as that from KVP40 can provide phylogenetically distant and tractable enzymes for further study of energy coupling. In our preliminary analysis, KVP40 NadV does show ATPase activity, and interestingly, the pZL405nadV ϩ clone complements the Salmonella Typhimurium pyridine nucleotide auxotroph even in the absence of promoter induction and noticeable enzyme synthesis ( Fig.…”

Section: Figmentioning

confidence: 99%

“…With ATP added, production of ADP was also detected, although the configuration of the assay did not allow detecting whether the reaction rate is higher in the presence of ATP. The KVP40 NadV NAmPRTase is therefore active, possessing properties similar to those of other nicotinate/nicotinamide phosphoribosyltransferases, including the facultative ATPase activity (16). NMNATase activity of NatV-His6.…”

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confidence: 93%

Vibrio Phage KVP40 Encodes a Functional NAD ⁺ Salvage Pathway

Lee

Miller

2017

J Bacteriol

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The genome of T4-type Vibrio bacteriophage KVP40 has five genes predicted to encode proteins of pyridine nucleotide metabolism, of which two, nadV and natV, would suffice for an NAD ϩ salvage pathway. NadV is an apparent nicotinamide phosphoribosyltransferase (NAmPRTase), and NatV is an apparent bifunctional nicotinamide mononucleotide adenylyltransferase (NMNATase) and nicotinamideadenine dinucleotide pyrophosphatase (Nudix hydrolase). Genes encoding the predicted salvage pathway were cloned and expressed in Escherichia coli, the proteins were purified, and their enzymatic properties were examined. KVP40 NadV NAmPRTase is active in vitro, and a clone complements a Salmonella mutant defective in both the bacterial de novo and salvage pathways. Similar to other NAmPRTases, the KVP40 enzyme displayed ATPase activity indicative of energy coupling in the reaction mechanism. The NatV NMNATase activity was measured in a coupled reaction system demonstrating NAD ϩ biosynthesis from nicotinamide, phosphoribosyl pyrophosphate, and ATP. The NatV Nudix hydrolase domain was also shown to be active, with preferred substrates of ADP-ribose, NAD ϩ , and NADH. Expression analysis using reverse transcriptionquantitative PCR (qRT-PCR) and enzyme assays of infected Vibrio parahaemolyticus cells demonstrated nadV and natV transcription during the early and delayed-early periods of infection when other KVP40 genes of nucleotide precursor metabolism are expressed. The distribution and phylogeny of NadV and NatV proteins among several large double-stranded DNA (dsDNA) myophages, and also those from some very large siphophages, suggest broad relevance of pyridine nucleotide scavenging in virus-infected cells. NAD ϩ biosynthesis presents another important metabolic resource control point by large, rapidly replicating dsDNA bacteriophages.IMPORTANCE T4-type bacteriophages enhance DNA precursor synthesis through reductive reactions that use NADH/NADPH as the electron donor and NAD ϩ for ADPribosylation of proteins involved in transcribing and translating the phage genome. We show here that phage KVP40 encodes a functional pyridine nucleotide scavenging pathway that is expressed during the metabolic period of the infection cycle. The pathway is conserved in other large, dsDNA phages in which the two genes, nadV and natV, share an evolutionary history in their respective phage-host group.

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Kinetic Mechanism of Nicotinic Acid Phosphoribosyltransferase: Implications for Energy Coupling

Cited by 34 publications

References 24 publications

Genomics-driven Reconstruction of Acinetobacter NAD Metabolism

Genomics-driven Reconstruction of Acinetobacter NAD Metabolism

Phosphoribosyl Diphosphate (PRPP): Biosynthesis, Enzymology, Utilization, and Metabolic Significance

Vibrio Phage KVP40 Encodes a Functional NAD ⁺ Salvage Pathway

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Kinetic Mechanism of Nicotinic Acid Phosphoribosyltransferase: Implications for Energy Coupling

Cited by 34 publications

References 24 publications

Genomics-driven Reconstruction of Acinetobacter NAD Metabolism

Genomics-driven Reconstruction of Acinetobacter NAD Metabolism

Phosphoribosyl Diphosphate (PRPP): Biosynthesis, Enzymology, Utilization, and Metabolic Significance

Vibrio Phage KVP40 Encodes a Functional NAD + Salvage Pathway

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Vibrio Phage KVP40 Encodes a Functional NAD ⁺ Salvage Pathway