Subunit location and sequences of the cysteinyl peptides of pig heart NAD-dependent isocitrate dehydrogenase.
Pig heart NAD-dependent isocitrate dehydrogenase has a subunit structure consisting of alpha 2 beta gamma, with the alpha subunit exhibiting a molecular weight of 39,000 and the beta and gamma each having molecular weights of 41,000. The amino-terminal sequences (33-35 residues) and the cysteinyl peptide sequences have now been determined by using subunits separated by chromatofocusing or isoelectric focusing and electroblotting. Displacement of the N-terminal sequence of the alpha subunit by 11-12 amino acids relative to that of the larger beta and gamma subunits reveals a 17 amino acid region of great similarity in which 10 residues are identical in all three subunits. The complete enzyme has 6.0 free SH groups per average subunit of 40,000 daltons, but yields 15 distinguishable cysteines in isolated tryptic peptides. Six distinct cysteines in sequenced peptides have been located in the alpha subunit. The beta and gamma subunits contain seven and five cysteines, respectively, with tryptic peptides containing three cysteines being common to the beta and gamma subunits. The three subunits appear to be closely related, but beta and gamma are more similar to each other than either is to the alpha subunit. The NAD-specific isocitrate dehydrogenase from pig heart has been shown to have 2 binding sites/enzyme tetramer for isocitrate, manganous ion, NAD+, and the allosteric activator ADP [Colman, R. F. (1983) Pept. Protein Rev. 1, 41-69]. It is proposed that the catalytically active tetrameric enzyme is organized as a dimer of dimers in which the alpha beta and alpha gamma dimers are nonidentical but functionally similar.
Huang
Y C
YC
Department of Chemistry and Biochemistry, University of Delaware, Newark 19716.
Colman
R F
RF
D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D003545:Cysteine; D007521:Isocitrate Dehydrogenase / Q000737:chemistry* / Q000235:genetics; D007525:Isoelectric Focusing; D008969:Molecular Sequence Data; D010088:Oxidoreductases / Q000235:genetics; D010446:Peptide Fragments / Q000302:isolation & purification; D010449:Peptide Mapping; D012689:Sequence Homology, Nucleic Acid; D013552:Swine
D010446:Peptide Fragments; D010088:Oxidoreductases; D007521:Isocitrate Dehydrogenase; D003545:Cysteine