Elevated Levels of Ketopantoate Hydroxymethyltransferase (PanB) Lead to a Physiologically Significant Coenzyme A Elevation in
            <i>Salmonella enterica</i>
            Serovar Typhimurium

Rubio, Aileen; Downs, Diana M.

doi:10.1128/jb.184.10.2827-2832.2002

Cited by 22 publications

(15 citation statements)

References 36 publications

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“…The thiamine requirement of each strain suppressed by the rpoD1181 allele could be satisfied by pantothenate (D. M. Downs, unpublished data). Past work showed that in some cases, the effect of pantothenate on thiamine synthesis was via an increase in CoA levels (21). Taken together, these results suggested that the rpoD mutation could be altering endogenous CoA levels to allow growth, possibly by increasing expression of the panBCD operon (Table 1).…”

Section: Vol 186 2004 Notes 4035supporting

confidence: 53%

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A Mutant Allele of rpoD Results in Increased Conversion of Aminoimidazole Ribotide to Hydroxymethyl Pyrimidine in Salmonella enterica

Dougherty

Downs

2004

J Bacteriol

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An allele of rpoD (rpoD1181) that results in increased synthesis of the pyrimidine moiety of thiamine in Salmonella enterica was identified. The S508Y substitution caused by rpoD1181 is analogous to the S506F derivative of the Escherichia coli protein. The properties of this E. coli mutant protein have been well characterized in vitro. Identification of a metabolic phenotype caused by the rpoD1181 allele of S. enterica allows past in vitro results to be incorporated in continuing efforts to understand cellular processes that are integrated with the thiamine biosynthetic pathway.The essential cofactor thiamine pyrophosphate is generated by the condensation of two independently synthesized molecules, 4-methyl-5(␤-hydroxyethyl) thiazole phosphate and 4-amino-5-hydroxymethyl-2-methylpyrimidine pyrophosphate (HMP-PP) ( Fig. 1) (3). The precursor of the pyrimidine moiety is 5-aminoimidazole ribotide (AIR), an intermediate in the purine biosynthetic pathway. HMP-P synthesis involves a complex intramolecular rearrangement and requires at least the product of the thiC gene (24, 26). Reconstitution of HMP-P synthesis in vitro has not yet been reported.Past work has identified loci other than thi biosynthetic enzymes that are required for efficient HMP-P synthesis. Characterization of these loci has implicated the oxidative pentose phosphate pathway, pantothenate (and/or coenzyme A [CoA]), and iron-sulfur cluster metabolism in the synthesis of HMP (10, 13, 14, 21, 23).Work described here was initiated to probe the integration of distinct metabolic processes with thiamine synthesis. Genetic analysis identified an allele of rpoD (rpoD1181) that allowed several HMP-requiring mutant strains to grow in the absence of thiamine. Our working model suggests that the rpoD1181 (S508Y) allele restores thiamine-independent growth by altering expression of an unidentified gene(s) whose product is required for efficient HMP-P synthesis.Thiamine synthesis in a purF iscA strain is restored by an rpoD allele. Salmonella enterica strain DM6176 (purF iscA) is unable to grow in the absence of thiamine (23). The iscA allele used is an insertion that is polar on hscAB (23). Mutations that suppressed the thiamine synthetic defect caused by the iscA lesion were identified. A 0.1-ml aliquot of an overnight culture of DM6176 was spread on a plate containing minimal medium plus gluconate and adenine. After overnight incubation at 37°C, more than 300 colonies that no longer required thiamine for growth were found. Ten random colonies were chosen for further analysis. In nine of the mutants, an insertion near the gltA locus, zbg-6391::Tn10d(Tc), was genetically linked to the causative lesion. On the basis of the proximity of gltA to the sdh genes and the knowledge that null mutations in sdh spared cellular thiamine pools (11), succinate dehydrogenase activity was measured in the nine mutants. Each of the nine mutants lacked detectable succinate dehydrogenase activity (data not shown), and they were not analyzed further.Multiple transposon insertions (M...

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Section: Vol 186 2004 Notes 4035supporting

confidence: 53%

“…Characterization of these loci has implicated the oxidative pentose phosphate pathway, pantothenate (and/or coenzyme A [CoA]), and iron-sulfur cluster metabolism in the synthesis of HMP (10,13,14,21,23).…”

mentioning

confidence: 99%

A Mutant Allele of rpoD Results in Increased Conversion of Aminoimidazole Ribotide to Hydroxymethyl Pyrimidine in Salmonella enterica

Dougherty

Downs

2004

J Bacteriol

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“…3C). This latter result narrowed the site of the previously described effect of coenzyme A on thiamine synthesis (17,24,43) to a role in the conversion of AIR to HMP, possibly due to its structural similarity to AIR.…”

Section: Thimentioning

confidence: 56%

Metabolic Flux in Both the Purine Mononucleotide and Histidine Biosynthetic Pathways Can Influence Synthesis of the Hydroxymethyl Pyrimidine Moiety of Thiamine in Salmonella enterica

2002

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Together, the biosyntheses of histidine, purines, and thiamine pyrophosphate (TPP) contain examples of convergent, divergent, and regulatory pathway integration. Mutations in two purine biosynthetic genes (purI and purH) affect TPP biosynthesis due to flux through the purine and histidine pathways. The molecular genetic characterization of purI mutants and their respective pseudorevertants resulted in the conclusion that <1% of the wild-type activity of the PurI enzyme was sufficient for thiamine but not for purine synthesis. The respective pseudorevertants were found to be informational suppressors. In addition, it was shown that accumulation of the purine intermediate aminoimidazole carboxamide ribotide inhibits thiamine synthesis, specifically affecting the conversion of aminoimidazole ribotide to hydroxymethyl pyrimidine.Cellular metabolism requires the control of flux through a complex network of pathways. This control is accomplished by regulation at several levels. Regulation of gene transcription or mRNA translation can control individual pathways. Allosteric control of the enzyme catalyzing the first committed step of a pathway is another mechanism used by cells to regulate biosynthetic pathways. Regulatory effects resulting from interactions between pathways are less well understood, despite the fact that many metabolites are present in multiple pathways in the cell.The expanding network of defined pathways that affect the biosynthesis of thiamine provides an attractive model system to investigate metabolic integration. As depicted in Fig. 1, the biosynthetic pathway for the hydroxymethylpyrimidine (HMP) moiety of thiamine pyrophosphate (TPP) branches off from that for purine mononucleotides at the metabolite aminoimidazole ribotide (AIR). Relevant to the work presented here is a subsequent purine intermediate, aminoimidazole carboxamide ribotide (AICAR), that is also a by-product of the biosynthesis of histidine.The distribution of AIR at the purine-HMP branch point must account for the different level of purine mononucleotides and TPP required for growth (purines/TPP ratio, 1,000:1, based on auxotrophic requirements). The distribution of AIR between these pathways could be accomplished by kinetic parameters of the relevant enzymes; however, the first dedicated HMP biosynthetic enzyme is poorly understood, thus hampering direct tests of this possibility.While thiamine and purine mononucleotides are synthesized by pathways diverging at the metabolite AIR, the cellular pool of AICAR arises from both purine and histidine biosynthesis and is used exclusively for purine mononucleotide synthesis (28, 29). Since AICAR enters purine mononucleotide synthesis after the purine-HMP branch point, no interaction between AICAR and HMP synthesis was anticipated. Hence, it was surprising to find that mutants blocked in the utilization of AICAR (purH mutants) required both purines and thiamine to grow (16, 58). The hypothesis proposed to explain this requirement posited that inactivation of purH would result in the acc...

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“…Alternatively, we hypothesize that a combination of Pp-coaA with enhanced pantothenate biosynthesis may also elevate CoA content without requiring an exogenous supply of precursors. It has been reported that pantothenate biosynthesis is enhanced by overexpression of ketopantoate hydroxymethyltransferase (PanB) from Salmonella enterica serovar Typhimurium (Rubio and Downs, 2002), PanD from Corynebacterium glutamicum (Dusch et al, E. coli W3110 cells transformed with Pp-coaA in a low-copy number plasmid were aerobically grown at 30°C for 24 h in M9 minimal medium supplemented with 2% glucose, 0.01% thiamine, 25 µg/ml chloramphenicol, and pantothenate at the indicated concentration. Cellular CoASH (black bar), acetyl-CoA (open bar), and malonyl-CoA (gray bar) were estimated by acyl-CoA cycling.…”

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confidence: 99%

Prokaryotic type III pantothenate kinase enhances coenzyme A biosynthesis in <i>Escherichia coli</i>

Ogata

Chohnan

2015

J. Gen. Appl. Microbiol.

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Elevated Levels of Ketopantoate Hydroxymethyltransferase (PanB) Lead to a Physiologically Significant Coenzyme A Elevation in Salmonella enterica Serovar Typhimurium

Cited by 22 publications

References 36 publications

A Mutant Allele of rpoD Results in Increased Conversion of Aminoimidazole Ribotide to Hydroxymethyl Pyrimidine in Salmonella enterica

A Mutant Allele of rpoD Results in Increased Conversion of Aminoimidazole Ribotide to Hydroxymethyl Pyrimidine in Salmonella enterica

Metabolic Flux in Both the Purine Mononucleotide and Histidine Biosynthetic Pathways Can Influence Synthesis of the Hydroxymethyl Pyrimidine Moiety of Thiamine in Salmonella enterica

Prokaryotic type III pantothenate kinase enhances coenzyme A biosynthesis in <i>Escherichia coli</i>

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