The subunit assembly of bovine erythrocyte superoxide dismutase was studied through analysis of the effects of heat, removal of metal ion cofactors and disulfide bond reduction, upon the dissociation of the subunits. Investigations utilizing sodium dodecyl sulfate acrylamide gel electrophoresis and sedimentation equilibrium analysis demonstrated that the 2 polypeptide subunits of the protein are not covalently joined, but are associated through unusually strong noncovalent interactions. This absence of intersubunit disulfide bonds, in conjunction with the titration of a single free sulfhydryl group per subunit polypeptide chain, necessitates that the 3 half cystine residues per subunit exist as 1 residue of cystine, forming an intrasubunit disulfide bond, and 1 residue of cysteine. The specific half cystine residues participating in the disulfide linkage and contributing the free thiol function were identified. Analysis of the products of cyanogen bromide cleavage, with acrylamide gel electrophoresis, demonstrated that the intrasubunit disulfide links 1 half cystine residue in the sequence NH2 terminal to Met 115, to another half cystine residue in the sequence COOH terminal to Met 115. Though conventional procedures of enzymatic digestion, peptide isolation and peptide characterization, Cys 55 and Cys 144 are shown to participate in the intrasubunit disulfide linkage, and Cys 6 to exist as free cysteine. (48 references).
|Original language||English (US)|
|Number of pages||9|
|Journal||Journal of Biological Chemistry|
|State||Published - Dec 1 1974|
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology