Acetate kinase from Clostridium acetobutylicum: a highly specific enzyme that is actively transcribed during acidogenesis and solventogenesis.
Acetate kinase (ATP:phosphotransferase, EC 2.7.2.1) has been purified 294-fold from acid-producing cells of Clostridium acetobutylicum DSM 1731 to a specific activity of 1087 U mg-1 (ADP-forming direction). The dimeric enzyme consisted of subunits with a molecular mass of 43 kDa. The molecular mass of the native acetate kinase was in the range 87-94 kDa as judged by gel filtration and native gel electrophoresis. The enzyme showed high specificity for the substrates acetate and ATP, and maximal activity was obtained with Mn2+ as divalent cation. The presence of mercury compounds such as HgCl2 and p-hydroxymercuribenzoate resulted in an essential loss of activity. The apparent K(m) values of acetate, Mg-ATP, acetyl phosphate, and Mg-ADP were 73, 0.37, 0.58 and 0.71 mM. An activity-staining procedure for detection of acetate kinase in crude cell extracts after separation on native polyacrylamide gels was developed. A DNA fragment encoding 246 bp of the acetate kinase gene of C. acetobutylicum DSM 792 was cloned by a PCR-based approach. Northern blot analysis revealed transcription of the gene under acid- and solvent-producing conditions with no significant differences at the level of transcription.
Winzer
Klaus
K
Angewandte Mikrobiologie und Mykoiogie, UniversitätUlm, D-89069 Ulm, Germany.
Lorenz
Karin
K
Angewandte Mikrobiologie und Mykoiogie, UniversitätUlm, D-89069 Ulm, Germany.
DÜrre
Peter
P
Angewandte Mikrobiologie und Mykoiogie, UniversitätUlm, D-89069 Ulm, Germany.
D000084:Acetate Kinase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D000143:Acids / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D003013:Clostridium / Q000201:enzymology* / Q000235:genetics* / Q000378:metabolism; D019281:Dimerization; D007700:Kinetics; D008970:Molecular Weight; D009838:Oligodeoxyribonucleotides / Q000235:genetics; D012997:Solvents / Q000378:metabolism; D013379:Substrate Specificity; D014158:Transcription, Genetic
D000143:Acids; D009838:Oligodeoxyribonucleotides; D012997:Solvents; D000084:Acetate Kinase