A method has been developed for the continuous spectrophotometric assay of dihydroxy-acetone synthase present in methanol-grown yeasts. This enzyme catalyses the condensation of formaldehyde with a pentose phosphate (most probably xylulose 5-phosphate) to give dihydroxyacetone and glyceraldehyde 3-phosphate. The assay is based on the NADH-and ATP-linked formation of L-glycerol 3-phosphate from dihydroxyacetone via the added coupling enzymes glycerol kinase and glycerol-3-phosphate dehydrogenase, and the reaction was followed by the decrease in absorbance at 340 nm. Using this assay, the pH optimum of the dihydroxyacetone synthase was shown to be 7.4 to 7.6. The enzyme, which catalyses a transketolase-like reaction, is a separate enzyme from the classical transketolase because their specific activities varied independently in and grown on different substrates and the two enzyme activities could be separated by ion-exchange chromatography.


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