Primary structure of the cholesterol side-chain cleavage cytochrome P-450 from bovine adrenocortical mitochondria and some aspects of its functioning on a structural level.
The primary structure of the cholesterol side-chain cleavage cytochrome P-450 (P-450scc) from bovine adrenocortical mitochondria has been determined. At the initial stage an exhaustive chymotryptic digestion of carboxymethylated P-450scc was performed, and the amino acid sequence of 66 peptides was determined. At the second stage an investigation of the amino acid sequence of individual fragments I (Mr 29 800) and II (Mr 26 600) of the limited trypsinolysis of P-450scc was carried out. Fragment I was digested with trypsin, Staphylococcus aureus V8 proteinase and thermolysin; fragment II was cleaved with trypsin and S. aureus V8 proteinase. In addition, the amino acid sequence of some CNBr peptides of P-450scc has been investigated. The primary structure of cytochrome P-450scc determined with protein chemistry methods proved the multistage cholesterol transformation to pregnenolone to be catalyzed by a single species of cytochrome P-450scc which consists of 481 amino acids. The results from protein sequencing of P-450scc are in good agreement with those obtained recently from nucleotide sequencing. The localization of peptide bonds cleaved under limited proteolysis of P-450 with trypsin to fragments I and II, I and III (Mr 16 800) is presented. It is shown that the transformation of P-450scc to P-420 is accompanied by the appearance of an additional trypsin-sensitive peptide bond in the N-terminal part of P-450scc.
D000302:Adrenal Cortex / Q000201:enzymology*; D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002845:Chromatography; D002918:Chymotrypsin; D003488:Cyanogen Bromide; D003577:Cytochrome P-450 Enzyme System; D010450:Endopeptidases; D008928:Mitochondria / Q000201:enzymology*; D010446:Peptide Fragments; D012697:Serine Endopeptidases; D013329:Structure-Activity Relationship; D014357:Trypsin
D010446:Peptide Fragments; D003577:Cytochrome P-450 Enzyme System; D010450:Endopeptidases; D012697:Serine Endopeptidases; D002918:Chymotrypsin; C028344:glutamyl endopeptidase; D014357:Trypsin; D003488:Cyanogen Bromide