Regulation of Clostridium acetobutylicum metabolism as revealed by mixed-substrate steady-state continuous cultures: role of NADH/NAD ratio and ATP pool.
Glycerol-glucose-fed (molar ratio of 2) chemostat cultures of Clostridium acetobutylicum were glucose limited but glycerol sufficient and had a high intracellular NADH/NAD ratio (I. Vasconcelos, L. Girbal, and P. Soucaille, J. Bacteriol. 176:1443-1450, 1994). We report here that the glyceraldehyde-3-phosphate dehydrogenase, one of the key enzymes of the glycolytic pathway, is inhibited by high NADH/NAD ratios. Partial substitution of glucose by pyruvate while maintaining glycerol concentration at a constant level allowed a higher consumption of glycerol in steady-state continuous cultures. However, glycerol-sufficient cultures had a constant flux through the glyceraldehyde-3-phosphate dehydrogenase and a constant NADH/NAD ratio. A high substitution of glucose by pyruvate [P/(G+P) value of 0.67 g/g] provided a carbon-limited culture with butanol and butyrate as the major end products. In this alcohologenic culture, the induction of the NADH-dependent butyraldehyde and the ferredoxin-NAD(P) reductases and the higher expression of alcohol dehydrogenases were related to a high NADH/NAD ratio and a low intracellular ATP concentration. In three different steady-state cultures, the in vitro phosphotransbutyrylase and butyrate-kinase activities decreased with the intracellular ATP concentration, suggesting a transcriptional regulation of these two genes, which are arranged in an operon (K. A. Walter, R. V. Nair, R. V. Carry, G. N. Bennett, and E. T. Papoutsakis, Gene 134:107-111, 1993).
Girbal
L
L
Centre de Bioingenierie Gilbert Durand, UA-CNRS no. 544, Institut National des Sciences Appliquées, Toulouse, France.
Soucaille
P
P
D000143:Acids / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000440:Butanols / Q000378:metabolism; D003013:Clostridium / Q000254:growth & development / Q000378:metabolism*; D000431:Ethanol / Q000378:metabolism; D005285:Fermentation; D005947:Glucose / Q000378:metabolism; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism*; D005990:Glycerol / Q000378:metabolism*; D006864:Hydrogenase / Q000378:metabolism; D008954:Models, Biological; D009243:NAD / Q000378:metabolism*; D009711:Nucleotides / Q000032:analysis; D010084:Oxidation-Reduction; D011773:Pyruvates / Q000378:metabolism; D013401:Sugar Alcohol Dehydrogenases / Q000378:metabolism
D000143:Acids; D000440:Butanols; D009711:Nucleotides; D011773:Pyruvates; D009243:NAD; D000431:Ethanol; D000255:Adenosine Triphosphate; D013401:Sugar Alcohol Dehydrogenases; C020672:glycerol dehydrogenase; D006864:Hydrogenase; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D005947:Glucose; D005990:Glycerol