Fine structure and biochemical properties of purified cytochrome oxidase

For the purpose to reveal the correlation between molecular structure and biochemical functions of cytochrome oxidase the author studied purified cytochrome oxidase by using high resolution electron microscope and biochemical methods. 1. Cytochrome oxidase was purified from the cytochrome oxidase-rich submitochondrial membrane (green membrane), obtained from beef heart mitochondria, by three different methods; modification of the method of OKUNUKI et ai., method of FOWLER et ai. and modification of the method ofJACOBS et ai. All the preparations showed a high specific activity under appropriate conditions and consisted mainly of small particles measuring approximately 80 to 90 A. in diameter. 2. The particle, measuring approximately 80 to 90 A. in diameter, took a cylindrical form measuring about 70 A. in diameter at the base and 95 A. in height in an appropriate condition. Many experimental results indicate that the particle is the smallest, fundamental unit of the active cytochrome oxidase. For this reason it was designated as the unit particle of cytochrome oxidase (abbreviated as UPCO). 3. The molecular weight of the unit particle, calculated from its volume and average density (1.24) of lipoproteins (3: 7), was about 270,000. The value was roughly twice the minimum molecular weight of 128, 000 calculated from the heme a content. Accordingly, it is considered that the unit particle contains two heme a molecule and two copper atoms. 4. It was suggested electron microscopically that the particle collected in the 22.6 S position by sucrose gradient ultracentrifugal analysis was a dimer of the unit particle of cytochrome oxidase and also that the particle collected in the 5. 7 S position was a half of the unit particle of cytochrome oxidase. 5. It was also suggested that the particle observed on the green membrane was a subunit of cytochrome oxidase, containing one heme a and one copper atom, and the unit particle of cytochrome oxidase was constituted of two of the particles observed on the green membrane. Namely, the results indicate that the molecular state of cytochrome oxidase on the green membrane apparently differs from that of the purified cytochrome oxidase.