Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0348321 (Haemophilus)
 15,372 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Our laboratory has previously reported a structurally and mechanistically related family of beta-hydroxyacid dehydrogenases with significant homology to beta-hydroxyisobutyrate dehydrogenase. A large number of the members of this family are hypothetical proteins of bacterial origin with unknown identity in terms of their substrate specificities and metabolic roles. The Escherichia coli beta-hydroxyacid dehydrogenase homologue corresponding to the locus was cloned and expressed with a 6-histidine tag for specific purification. The purified recombinant protein very specifically catalyzed the NAD(+)-dependent oxidation of d-glycerate and the NADH-dependent reduction of tartronate semialdehyde, identifying this protein as a tartronate semialdehyde reductase. Further evidence for identification as tartronate semialdehyde reductase is the observation that the coding region for this protein is directly preceded by genes coding for hydroxypyruvate isomerase and glyoxylate carboligase, two enzymes that synthesize tartronate semialdehyde, producing an operon clearly designed for d-glycerate biosynthesis from tartronate semialdehyde. The single beta-hydroxyacid dehydrogenase homologue from Haemophilus influenzae was also cloned, expressed, and purified with a 6-histidine tag. This protein also catalyzed the NAD(+)-dependent oxidation of d-glycerate but was significantly more efficient in the oxidation of four-carbon beta-hydroxyacids like d-hydroxybutyrate and d-threonine. This enzyme differs from all the presently known beta-hydroxybutyrate dehydrogenases which are well established members of the short chain dehydrogenase/reductase superfamily.
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PMID:Novel beta -hydroxyacid dehydrogenases in Escherichia coli and Haemophilus influenzae. 1097 49

CDP-ribitol synthase is a bifunctional reductase and cytidylyltransferase that catalyzes the transformation of D-ribulose 5-phosphate, NADPH, and CTP to CDP-ribitol, a repeating unit present in the virulence-associated polysaccharide capsules of Haemophilus influenzae types a and b [Follens, A., et al. (1999) J. Bacteriol. 181, 2001]. In the work described here, we investigated the order of the reactions catalyzed by CDP-ribitol synthase and conducted experiments to resolve the question of substrate channeling in this bifunctional enzyme. It was determined that the synthase first catalyzed the reduction of D-ribulose 5-phosphate followed by cytidylyl transfer to D-ribitol 5-phosphate. Steady state kinetic measurements revealed a 650-fold kinetic preference for cytidylyl transfer to D-ribitol 5-phosphate over D-ribulose 5-phosphate. Rapid mixing studies indicated quick reduction of D-ribulose 5-phosphate with a lag in the cytidylyl transfer reaction, consistent with a requirement for the accumulation of K(m) quantities of D-ribitol 5-phosphate. Signature motifs in the C-terminal and N-terminal sequences of the enzyme (short chain dehydrogenase/reductase and nucleotidyltransferase motifs, respectively) were targeted with site-directed mutagenesis to generate variants that were impaired for only one of the two activities (K386A and R18A impaired for reduction and cytidylyl transfer, respectively). Release and free diffusion of the metabolic intermediate D-ribitol 5-phosphate was indicated by the finding that equimolar mixtures of K386A and R18A variants were efficient for bifunctional catalysis. Taken together, these findings suggest that bifunctional turnover occurs in distinct active sites of CDP-ribitol synthase with reduction of D-ribulose 5-phosphate and release and free diffusion of the metabolic intermediate D-ribitol 5-phosphate followed by cytidylyl transfer.
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PMID:Reduction precedes cytidylyl transfer without substrate channeling in distinct active sites of the bifunctional CDP-ribitol synthase from Haemophilus influenzae. 1130 20