Glucose repression and hexokinase isoenzymes in yeast. Isolation and characterization of a modified hexokinase PII isoenzyme.
Hexokinase PII, but not isoenzyme PI, has a unique role in glucose repression in yeasts [Entian, K.-D. (1980) Mol. Gen. Genet. 178, 633-637; Entian, K.-D. and Mecke, D. (1982) J. Biol. Chem. 257, 870-874; Entian, K.-D. and Fröhlich, K.-U. (1984) J. Bacteriol. 158, 29-35]. The number of hexokinase isoenzymes in crude extracts was re-examined by chromatofocusing. In addition to the known isoenzymes PI and PII, a third isoenzyme, PIIM, was detected. The activity of this enzyme was only about 5-10% of that of hexokinase PII and was independent of growth conditions. Experiments with hexokinase transformants and purified hexokinase isoenzymes clearly indicated that the PIIM form is also present in vivo. Fingerprint mapping of purified hexokinases showed that hexokinase PIIM is closely related to PII. Hybridization experiments between totally restricted yeast DNA and the previously isolated PII gene clearly indicated that PIIM is also coded by one of the two known hexokinase genes. No mRNA specific for hexokinase PIIM was detected after hybridization experiments with the previously cloned hexokinase PII gene [Fröhlich et al. (1984) Mol. Gen. Genet. 194, 144-148]. Hexokinase PIIM appears to be derived from hexokinase PII by a posttranslational event. The Km values of each of the purified isoenzymes, PII and PIIM, were identical for glucose, fructose and ATP. Both isoenzymes were strongly inhibited by high physiological concentrations for ATP; such inhibition has not been described previously. The possible role of hexokinase PIIM in glucose repression is discussed.
D055598:Chemical Phenomena; D002621:Chemistry; D002845:Chromatography / Q000379:methods; D004271:DNA, Fungal; D005947:Glucose / Q000494:pharmacology*; D006593:Hexokinase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification*; D006358:Hot Temperature; D007525:Isoelectric Focusing; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification*; D009154:Mutation; D009693:Nucleic Acid Hybridization; D012331:RNA, Fungal; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics
D004271:DNA, Fungal; D007527:Isoenzymes; D012331:RNA, Fungal; D006593:Hexokinase; D005947:Glucose