Levels of key enzymes and physiological factors involved in product formation in batch and continuous cultures of Clostridium acetobutylicum ATCC 824

[摘要] A number of physiological factors and their relation to the solventogenesis mechanism were investigated in both batch and continuous cultures of Clostridium acetobutylicum. The initiation of butanol formation in batch culture correlates best with minimum levels of undissociated butyric acid in the medium. Weak organic acids have uncoupling characteristics and effect a concentration-dependent decrease in the transmembrane $Delta$pH and the intracellular pH (pH$sb{i}$). The pH$sb{i}$ decreases continuously from 7.0 to 5.5 in batch cultures and cell growth terminates when the pH$sb{i}$ drops below 5.5.Product selectivities in continuous culture are dependent on the supply and demand of biosynthetic energy. Glucose limitation leads to ATP-limited culture conditions and acid formation. By contrast, glucose-sufficient fermentations are characterized by high intracellular ATP concentrations and elevated solvent yields. Carbon monoxide gassing of an acidogenic continuous culture leads to increased levels of intracellular ATP and NAD(P)H concentrations as well as elevated pH$sb i$ and $Delta$pH values.The formation of butanol in continuous culture is primarily regulated at the genetic level via expression of NADP-specific butyraldehyde dehydrogenase (BYDH (NADP)). Addition of glucose, butyric acid and carbon monoxide results in induction of this enzyme. In contrast to continuous culture, in vitro activities of BYDH (NADP) do not correlate with solventogenesis in controlled pH batch fermentations.The production of acetone in continuous fermentations is also controlled at the genetic level through expression of CoA-transferase (COA-T). This enzyme is induced by glucose. Acetoacetate decarboxylase (ACACDE) is inactivated by carbon monoxide. The production of acetone in controlled pH batch fermentations is regulated at the genetic level through the combined induction of both CoA-transferase and acetoacetate decarboxylase. Neither in vitro activities of COA-T and ACACDE nor acetone formation could be detected above pH 6.0 in either batch or continuous culture.The formation of organic acids by Clostridium acetobutylicum appears to be controlled at the enzyme level. Several possible control mechanisms operating at the enzyme level are discussed. Finally, the uptake of butyrate is not necessarily linked to acetone formation and can proceed via a reversal of the phosphotransbutyrylase and butyrate kinase reaction pathway.

[发布机构] Rice University

[效力级别] engineering