Purification and characterization of formaldehyde dehydrogenase from rat liver cytosol.
Formaldehyde dehydrogenase was purified to electrophoretic and column chromatographic homogeneity from rat liver cytosolic fraction by a procedure which includes ammonium sulfate precipitation, DEAE-cellulose-, hydroxyapatite-, Mono Q-chromatography, and gel filtration. Its molecular mass was estimated to be 41 kDa by gel filtration and SDS-PAGE, suggesting that it is a monomer. It utilized neither methylglyoxal nor aldehydes except formaldehyde as a substrate. It has been reported that liver class III alcohol dehydrogenase and formaldehyde dehydrogenase are the same enzyme and oxidize formaldehyde and long chain primary alcohols. However, the enzyme examined here did not use n-octanoi as a substrate. The Km values for formaldehyde and NAD+ were 5.09 and 2.34 microM at 25 degrees C, respectively. The amino acid sequences of 10 peptides obtained from the purified enzyme after digestion with either V8 protease or lysyl endopeptidase were determined. From these results, the enzyme was proved to be different from the previously described mammalian formaldehyde dehydrogenase and is the first true formaldehyde dehydrogenase to be isolated from a mammalian source.
Tsuboi
S
S
Faculty of Pharmaceutical Sciences, Okayama University.
Kawase
M
M
Takada
A
A
Hiramatsu
M
M
Wada
Y
Y
Kawakami
Y
Y
Ikeda
M
M
Ohmori
S
S
D000445:Aldehyde Oxidoreductases / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000502:physiology; D000595:Amino Acid Sequence; D000818:Animals; D003600:Cytosol / Q000201:enzymology*; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D007700:Kinetics; D008099:Liver / Q000201:enzymology* / Q000648:ultrastructure; D008297:Male; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000032:analysis / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D012697:Serine Endopeptidases / Q000378:metabolism; D013379:Substrate Specificity
D010446:Peptide Fragments; D000445:Aldehyde Oxidoreductases; C087006:formaldehyde dehydrogenase, glutathione-independent; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; C036813:lysyl endopeptidase