Effect of specific trifluoroacetylation of individual cytochrome c lysines on the reaction with cytochrome oxidase.
We have prepared three different cytochrome c derivatives, each containing a single specifically trifluoroacetylated lysine at residues 13, 55, and 99, respectively. The only modification that affected cytochrome c oxidase (EC 1.9.3.1) activity was that of lysine-13 at the top of the heme crevice. Trifluoroacetylation of lysine-13 increased the apparent Michaelis constant fivefold compared to that of native cytochrome c, but did not affect the maximum velocity. Trifluoroacetylation of lysine-55 at the left side of the cytochrome c molecule did not affect cytochrome oxidase activity in any way, nor did trifluoroacetylation of lysine-99 at the rear of the cytochrome c molecule. This indicates that the cytochrome oxidase binding site on cytochrome c involved only the front of the cytochrome c molecule and those lysines immediately surrounding the heme crevice.
D000818:Animals; D001665:Binding Sites; D003574:Cytochrome c Group; D003576:Electron Transport Complex IV / Q000378:metabolism; D006736:Horses; D007700:Kinetics; D008239:Lysine; D009682:Magnetic Resonance Spectroscopy; D009206:Myocardium / Q000201:enzymology; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000032:analysis; D011485:Protein Binding; D011487:Protein Conformation; D014269:Trifluoroacetic Acid; D014357:Trypsin
D003574:Cytochrome c Group; D010446:Peptide Fragments; D014269:Trifluoroacetic Acid; D003576:Electron Transport Complex IV; D014357:Trypsin; D008239:Lysine