Novel separation and amino acid sequences of alpha and beta subunits of pig heart pyruvate dehydrogenase.
Pyruvate dehydrogenase was separated from pig heart pyruvate dehydrogenase complex by gel-permeation chromatography on a TSK G4000SWG column in the presence of 4 M urea, followed by chromatography on a calcium phosphate gel-cellulose column. The pyruvate dehydrogenase was further separated into two nonidentical subunits, alpha and beta, by high-performance liquid chromatography on a Synchropak CM-300 column in the presence of 8 M urea. The complete amino acid sequences of two subunits of pyruvate dehydrogenase were determined. The peptide fragments of S-carboxymethylated subunits were generated by treatment with endoproteinase Lys-C, endoproteinase Asp-N, trypsin, and cyanogen bromide. The subunits alpha and beta contain 361 amino acid residues (Mr 40,294) and 329 residues (Mr 35,787), respectively. The amino acid sequences of subunits alpha and beta in the pig were, respectively, 98 and 96% identical to those in humans. Hydropathy analysis and prediction of the secondary structure of two subunits suggest that the subunit alpha contains the thiamin pyrophosphate-binding domain and that the subunit beta contains segments with a high hydrophobicity.
Urata
Y
Y
Department of Pathological Biochemistry, Nagasaki University School of Medicine, Japan.
Koike
K
K
Goto
S
S
Koike
M
M
D000595:Amino Acid Sequence; D000818:Animals; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D008969:Molecular Sequence Data; D009206:Myocardium / Q000737:chemistry*; D011768:Pyruvate Dehydrogenase Complex / Q000737:chemistry* / Q000302:isolation & purification; D013552:Swine
D011768:Pyruvate Dehydrogenase Complex