Immobilized cytochrome P-450LM2 : dissociation and reassociation of oligomers

• Published in: FEBS letters 1990-09, Vol.270 (1-2), p.177-180
• Main Authors: MYASOEDOVA, K. N, BERNDT, P
• Format: Article
• Language: English
• Subjects: Analytical, structural and metabolic biochemistry; Animals; Biological and medical sciences; Cholic Acid; Cholic Acids; Cytochrome P-450 Enzyme System - chemistry; Detergents; Dextrans; Enzymes and enzyme inhibitors; Enzymes, Immobilized - chemistry; Fundamental and applied biological sciences. Psychology; Glucosides; Hydrogen-Ion Concentration; Kinetics; Macromolecular Substances; Oxidoreductases; Phospholipids; Polyethylene Glycols; Rabbits; Sepharose; Sodium Dodecyl Sulfate; Temperature
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/0014-5793(90)81261-L

Subunit interactions in the purified hexameric cytochrome P-450LM2 have been studied using covalent binding of one of the 6 promoters to an insoluble matrix. High ionic strength, large-scale pH changes, guanidine chloride and sodium cholate taken at membrane-solubilizing concentrations, had no effect on the aggregation state of the immobilized hemoprotein. SDS caused a 6-fold decrease in the amount of the bound cytochrome. Non-ionic detergents (Emulgen 913, octylglucoside, Tritons) induced hexamer dissociation. In the presence of Emulgen 913 (greater than 0.2%), monomers and immobilized dimers were obtained as cytochrome P-450 was studied in an aqueous medium and in the immobilized state, respectively. Immobilized dimers could be reconstituted to hexamers by treatment with an excess of solubilized monomers after removal of the detergent. In the presence of various phospholipids, which increased the immobilized cytochrome P-450LM2 demethylase activity and induced characteristic spectral changes, no hexamer dissociation was shown. The data obtained are thus in agreement with the suggestion that hexameric arrangement is inherent in the cytochrome P-450 when it is bound to the native membranes.