The glyceraldehyde-3-phosphate dehydrogenase of Clostridium acetobutylicum: isolation and purification of the enzyme, and sequencing and localization of the gap gene within a cluster of other glycolytic genes.
Glyceraldehyde-3-phosphate dehydrogenase was purified from Clostridium acetobutylicum by sequential ammonium sulfate precipitation, gel filtration and anion-exchange chromatography (to a specific activity of 27 U mg-1). The enzyme had a molecular mass of 40 kDa as determined by SDS-PAGE and a native molecular mass of 160 kDa as determined by nondenaturing PAGE, indicating that it has a homotetrameric composition. Its pH optimum was between 8.5 and 9.3. The corresponding gene (gap) was cloned and sequenced from C. acetobutylicum DSM 792 and found to cluster with other genes of enzymes from the glycolytic pathway (pgk, phosphoglycerate kinase; tpi, triosephosphate isomerase; pgm(i), 2,3-bisphosphoglycerate-independent phosphoglycerate mutase). No sequences resembling rho-independent transcription terminators were found in the intergenic regions. A plasmid carrying the clostridial gap gene complemented an Escherichia coli gap mutant.
Schreiber
Wiebke
W
Dürre
Peter
P
D003001:Cloning, Molecular; D003013:Clostridium / Q000201:enzymology* / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005798:Genes, Bacterial / Q000235:genetics*; D005816:Genetic Complementation Test; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism; D006019:Glycolysis; D008969:Molecular Sequence Data; D005810:Multigene Family / Q000235:genetics; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction / Q000379:methods; D017422:Sequence Analysis, DNA
D005987:Glyceraldehyde-3-Phosphate Dehydrogenases