Studies on cytochrome oxidase. Partial resolution of enzymes containing seven or six subunits, from yeast and beef heart, respectively.
Highly active, essentially homogeneous, preparations of ferrocytochrome c oxidase (EC 1.9.3.1) have been obtained from both yeast and beef heart by extraction with cholate, fractionation with ammonium sulfate, and replacement of cholate by Tween 20. The molecular weights of the resultant proteins equal 260 +/- 23 X 10(3) and 205 +/- 10(3); they contain seven and six different polypeptide subunits, respectively, all in equimolar amounts, with apparent molecular weights of 42.4, 34.1, 24.7, 14.6, 14.6, 12.3, 10.6 X 10(3), and 47.5, 20.4, 14.5, 14.5, 13.0, 11.0 X 10(3), respectively. By means of apolar chromatography on L-leucine coupled to agarose these enzymes can be stripped of their largest subunit(s) resulting in preparations with molecular weights of 170 +/- 17 X 10(3) and 124 +/- 20 X 10(3), and containing only five polypeptides, with the largest remaining one (molecular weight congruent to 20 X 10(3)) present in less than stoichiometric amounts. This interconversion and subunit removal has been monitored by exclusion chromatography, four systems of acrylamide gel electrophoresis--some with the protein labeled with 125I under denaturing conditions--isoelectric focusing, and hydrodynamic methods. It has virtually no effect on heme a and copper content and on the catalytic parameters of the enzymes.  We conclude that subunits I and II in enzymes from fungal, and subunit I in those from animal, sources are dispensable for the catalysis of the oxidation of ferrocytochrome c by, and are probably not essential for the attatchment of prosthetic groups to, these proteins.
D000818:Animals; D002417:Cattle; D003300:Copper / Q000032:analysis; D003576:Electron Transport Complex IV / Q000032:analysis* / Q000302:isolation & purification / Q000378:metabolism; D006418:Heme / Q000032:analysis; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D046911:Macromolecular Substances; D008928:Mitochondria / Q000201:enzymology*; D008931:Mitochondria, Muscle / Q000201:enzymology*; D008970:Molecular Weight; D009206:Myocardium; D010743:Phospholipids / Q000032:analysis; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013045:Species Specificity; D013056:Spectrophotometry, Ultraviolet
D046911:Macromolecular Substances; D010743:Phospholipids; D006418:Heme; D003300:Copper; D003576:Electron Transport Complex IV