Molecular and physiological aspects of alcohol dehydrogenases in the ethanol metabolism Saccharomyces cerevisiae

[摘要] English: When Saccharomyces cerevisiae is grown on a fermentable carbon source such as glucose, thefermentative alcohol dehydrogenase, ADH I , catalyses the regeneration of NAD+ from NADHand produces ethanol from acetaldehyde. When the fermentable carbon source is depleted, avariety of other enzymes are derepressed in order to utilise the previously excreted ethanol viaoxidative respiration and gluconeogenesis . To provide both the carbon source and energy forthis system, the yeast cell requires an efficient method for oxidising this previously excretedethanol. ADH II is a catabolite repressible isoenzyme which primarily functions in the cell tooxidise ethanol to acetaldehyde, which can be metabolised via the tricarboxylic acid cycle or actas intermediate product in gluconeogenesis. ADH III is a mitochondrial isoenzyme participatingin the respiratory metabolism by forming part of the ethanol-acetaldehyde shuttle that isimportant for shuttling mitochondrial reducing equivalents to the cytosol under anaerobicconditions. The physiological roles and regulation of ADH1, ADH2, ADH3, ADH4 and ADH5were investigated by monitoring transcription levels in chemostat and batch cultivations withNorthern blotting and real-time RT-PCR. ADH I was shown to be the key enzyme in thereduction of acetaldehyde to ethanol and also demonstrated ample ability to oxidise ethanol.ADH2 transcription was inhibited by glucose and ethanol in chemostat cultures pulsed with boththese carbon sources, but only glucose repression was evident in batch cultures. Northern blotanalysis showed that the ADH3 gene was induced during the ethanol phase of the pulsessuggested that the mitochondrial ADH III enzyme could also be involved in the first step inethanol utilisation. The growth kinetics of a strain expressing only ADH III demonstrated that theADH3 gene product could fulfil the same function as ADH II. ADH4 transcription was detectedfor the first time in batch cultures and was shown not to be involved in the production orassimilation of ethanol. ADH5 transcription was also demonstrated for the first time and datasuggest that ADH V is not involved in ethanol production in a adh1-adh4 deletion mutant.

[发布机构] University of the Free State