Alpha,beta-dicarbonyl reduction by Saccharomyces D-arabinose dehydrogenase.
An alpha,beta-dicarbonyl reductase activity was purified from Saccharomyces cerevisiae and identified as the cytosolic enzyme D-Arabinose dehydrogenase (ARA1) by MALDI-TOF/TOF. Size exclusion chromatography analysis of recombinant Ara1p revealed that this protein formed a homodimer. Ara1p catalyzed the reduction of the reactive alpha,beta-dicarbonyl compounds methylglyoxal, diacetyl, and pentanedione in a NADPH dependant manner. Ara1p had apparent Km values of approximately 14 mM, 7 mM and 4 mM for methylglyoxal, diacetyl and pentanedione respectively, with corresponding turnover rates of 4.4, 6.9 and 5.9 s(-1) at pH 7.0. pH profiling showed that Ara1p had a pH optimum of 4.5 for the diacetyl reduction reaction. Ara1p also catalyzed the NADP+ dependant oxidation of acetoin; however this back reaction only occurred at alkaline pH values. That Ara1p was important for degradation of alpha,beta-dicarbonyl substrates was further supported by the observation that ara1-Delta knockout yeast mutants exhibited a decreased growth rate phenotype in media containing diacetyl.
van Bergen
Barry
B
Department of Bioresource Engineering, Macdonald Campus of McGill University, 21 111 Lakeshore Road, Ste-Anne-de-Bellevue, Quebec, Canada H9X 3V9.
Strasser
Rona
R
Cyr
Normand
N
Sheppard
John D
JD
Jardim
Armando
A
D000094:Acetoin Dehydrogenase / Q000737:chemistry / Q000302:isolation & purification; D000595:Amino Acid Sequence; D003931:Diacetyl / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D050837:Oxidoreductases Acting on Aldehyde or Oxo Group Donors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011765:Pyruvaldehyde / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013401:Sugar Alcohol Dehydrogenases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
C000588781:Ara1 protein, S cerevisiae; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D011765:Pyruvaldehyde; D013401:Sugar Alcohol Dehydrogenases; D000094:Acetoin Dehydrogenase; C028590:D-arabinose dehydrogenase; D050837:Oxidoreductases Acting on Aldehyde or Oxo Group Donors; D003931:Diacetyl