title: ApbA, the ketopantoate reductase enzyme of Salmonella typhimurium is required for the synthesis of thiamine via the alternative pyrimidine biosynthetic pathway.
abstract: The apbA gene of Salmonella typhimurium was shown to encode ketopantoic acid reductase. ApbA was purified from crude cell-free extracts to greater than 95% homogeneity after two chromatographic steps. N-terminal amino acid sequencing (first 15 amino acids) and Western blot analysis confirmed the isolated protein was ApbA. The functional protein was a monomer with a molecular mass of 31.1 kDa. Optimal reaction conditions for the reduction of ketopantoic acid were established at a pH of 6.25, and a temperature of 42 degreesC. The preferred electron source was NADPH, and the apparent Km constants of the enzyme for NADPH and ketopantoic acid were determined to be 0.776 +/- 0.09 mM and 0.742 +/- 0.01 mM, respectively. The homogeneous enzyme had a specific activity of 64.3.
authors.affiliation: Department of Bacteriology, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA.
authors.forename: M E
authors.initials: ME
authors.lastname: Frodyma
authors.forename: D
authors.initials: D
authors.lastname: Downs
mesh_terms: D000429:Alcohol Oxidoreductases / Q000378:metabolism*; D002087:Butyrates / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009249:NADP / Q000378:metabolism; D011743:Pyrimidines / Q000096:biosynthesis*; D011994:Recombinant Proteins / Q000378:metabolism; D012486:Salmonella typhimurium / Q000201:enzymology*; D017421:Sequence Analysis; D013696:Temperature; D013831:Thiamine / Q000096:biosynthesis*
chemical_list: D002087:Butyrates; D011743:Pyrimidines; D011994:Recombinant Proteins; D009249:NADP; D000429:Alcohol Oxidoreductases; C021934:2-dehydropantoate 2-reductase; D013831:Thiamine