ISOLATION OF MITOCHONDRIAL SUCCINATE - UBIQUINONE REDUCTASE, CYTOCHROME-C REDUCTASE AND CYTOCHROME-C OXIDASE FROM NEUROSPORA-CRASSA USING NON-IONIC DETERGENT

The electron transfer complexes, succinate: ubiquinone reductase, ubiquinone: cytochrome c reductase, and cytochrome c:O2 oxidase were isolated from the mitochondrial membranes of Neurospora crassa by the following steps. Modification of the contents of the complexes in mitochondria by growing cells on chloramphenicol; solubilisation of the complexes by Triton X‐100; affinity chromatography on immobilized cytochrome c and ion exchange and gel chromatography. Ubiquinone reductase was obtained in a monomeric form (Mr∼ 130000) consisting of a flavin subunit (Mr 72000) an iron‐sulfur subunit (Mr 28000) and a cytochrome b subunit (Mr probably 14000). Cytochrome c reductase was obtained in a dimeric form (Mr∼ 550000), the monomeric unit comprising two cytochromes b (Mr each 30000), a cytochrome c1 (Mr 31000), an iron‐sulfur subunit (Mr 25000), and six subunits without known prosthetic groups (Mr 9000, 11000, 14000, 45000, 45000, and 52000). Cytochrome c oxidase was also isolated in a dimeric form (Mr∼ 320000) comprising two copies each of seven subunits (Mr 9000, 12000, 14000, 18000, 21000, 29000, and 40000). The complexes were essentially free of phospholipid. Each bound one micelle of Triton X‐100 (Mr∼ 90000). After isolation, the bound Triton X‐100 could be replaced by other non‐ionic detergents such as: alkylphenyl polyoxyethylene ethers, alkyl polyoxyethylene ethers and acyl polyoxyethylene sorbitan esters. Copyright © 1979, Wiley Blackwell. All rights reserved