Structure of the cytochrome c oxidase complex of rat liver. 1. Studies on nearest-neighbour relationship of polypeptides with cross-linking reagents.

Isolated rat liver cytochrome c oxidase was cross-linked with the cleavable reagents dimethyl-3,3'-dithiobispropionimidate (DTBP), 3,3'-dithiobis(succinimidyl)propionate (DSP) and cupric di(1,10-phenanthroline) (CuP). The cross-linked products were separated by high-resolving two-dimensional dodecyl sulfate gel electrophoresis, which separates all thirteen polypeptides of the mammalian enzyme. With cupric di(1,10-phenanthroline) seven polypeptides (I-III, Va, Vb, VIIb and VIII) were cross-linked with each other and with itself, indicating the occurrence of free -SH groups in these polypeptides and a rearrangement of the native structure of the complex by cupric di(1,10-phenanthroline). With dimethyl-3,3'-dithiobispropionimidate or 3,3'-dithiobis(succinimidyl)propionate all nuclear-coded polypeptides, with the exception of polypeptide VIIa, formed cross-linked products with the three 'catalytic' polypeptides I-III, which are coded on mitochondrial DNA. Five additional cross-linked pairs were found between nuclear coded polypeptides. The close arrangement of nuclear coded polypeptides with the catalytic polypeptides suggests a regulatory function of these polypeptides.