Summary: The pattern of ethanol and lactate formation by continuous cultures of LMG 6564 under glucose limitation is affected by culture conditions such as pH and dilution rate. NADH- and NADPH-mediated lactate dehydrogenase (LDH; EC and alcohol (ethanol) dehydrogenase (ADH; EC and EC activities were measured in cell extracts from continuous cultures grown under different conditions. In conditions of high product formation, the NADH-mediated reaction was higher than the NADPH-mediated reaction for both LDH and ADH. LDH showed an absolute requirement for fructose 1,6-bisphosphate (FBP). Both NADH- and NADPH-linked LDH reactions were cytoplasmic, not sensitive to oxygen, and had a pH optimum of 6.0–6.5; the temperature optimum was 55–60°C. The reverse reaction (lactate oxidation) could not be demonstrated. ADH activity was found in the particulate fraction of the cell lysate and was sensitive (not completely, but irreversibly) to oxygen. The temperature and pH optima were 43°C, pH 7.0 and 45°C, pH 8.8 for the NADH- and NADPH-mediated reactions, respectively. The production of at least two different ADHs is likely. LDH and ADH seemed to be regulated at the level of enzyme synthesis (direct correlation between the activities and the lactate and ethanol yields in the culture) with a second regulation of LDH by FBP at the reaction level.


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