Localization of the N-ethylmaleimide reactive cysteine in the beef heart mitochondrial ADP/ATP carrier protein.
Alkylation of the ADP/ATP carrier protein in beef heart mitochondria by N-ethylmaleimide (NEM) results in inactivation of transport. One out of the four cysteinyl residues contained in 1 mol of carrier subunit of Mr 32 000 is alkylated by NEM. The identification of the alkylated residue to Cys-56 has been achieved by chemical and enzymatic cleavages. The chemical cleavages included cleavages at the nonalkylated cysteinyl residues by cyanide at alkaline pH and at methionyl residues by cyanogen bromide. Enzymatic cleavage involved the use of trypsin and chymotrypsin; the resulting peptides were resolved by high-performance liquid chromatography. Analysis of a small size [14C]NEM-labeled peptide obtained by tryptic and chymotryptic digestion of the [14C]NEM-labeled carrier protein yielded the following amino acid sequence: (Formula: see text) where X is probably a substituted lysine.
D000478:Alkylation; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002417:Cattle; D003545:Cysteine / Q000032:analysis*; D005033:Ethylmaleimide; D008715:Methionine / Q000032:analysis; D008929:Mitochondria, Heart / Q000201:enzymology*; D000226:Mitochondrial ADP, ATP Translocases / Q000378:metabolism*; D009713:Nucleotidyltransferases / Q000378:metabolism*; D010446:Peptide Fragments / Q000032:analysis
D000596:Amino Acids; D010446:Peptide Fragments; D000226:Mitochondrial ADP, ATP Translocases; D008715:Methionine; D009713:Nucleotidyltransferases; D003545:Cysteine; D005033:Ethylmaleimide