The role of phospholipids in the reduction of ubiquinone analogues by the mitochondrial reduced nicotinamide-adenine dinucleotide-ubiquinone oxidoreductase complex

Abstract

The isolated NADH-ubiquinone oxidoreductase complex of bovine heart mitochondria reduces ubiquinone analogues by two pathways. One pathway is inhibited by rotenone, and reduction of quinones takes place in the lipid phase of the system. The other pathway is insensitive to rotenone and reduction takes place in the aqueous phase. The variation of rates of electron transpport with the chemical nature of the quinone analogue and the concentrations of both quinone and phospholipid can be rationalized in terms of partition of the quinone between the aqueous and lipid phases of the system. Thus one function of phospholipid associated with the enzyme appears to be to act as solvent for ubiquinone reduced by the rotenone-sensitive pathway. This proposal is supported by the kinetic behaviour of enzyme whose endogenous lipids have been replaced by (1,2)-dimyristoylsn-glycero-3-phosphocholine. Thus, under certain circumstances, the rotenone-sensitive reduction of ubiquinone-1 exhibited a substantial increase in activation energy below the phase-transition temperature of the synthetic lipid, whereas the reduction of other acceptors was unaffected.

The effects of lipid phase transitions on the interaction of mitochondrial NADH–ubiquinone oxidoreductase with ubiquinol–cytochrome c oxidoreductase

“…lOa), lowering the temperature to below the phase-transition temperature of the lipid had a relatively small effect on activity, the activation energy rising between 20 and 4OkJ/mol depending on conditions. When Complex I was assayed with exogenous ubiquinone-l as substrate, phase transition had no effect on activity unless a large excess of phospholipid was present (Ragan, 1978). Complex-III activity with ubiquinol-2 as substrate was not affetted by phase transitions whatever the lipid content.…”

“…Ragan, 1978), it was important to establish that the behaviour of the Complex I-IIl system was maintained after removal of endogenous detergents by dialysis. In the experiment of Table 3, presence or absence of 'Qpool' behaviour was assessed by the ratio of the NADH-cytochrome c oxidoreductase activity obtained at a low Complex-IlI concentration to that obtained at a high concentration .…”

“…These units dissociate and reassociate randomly at rates exceeding the electron-transfer rates, thereby mixing the associated and unassociated ubiquinone-10 and leading to so-called 'Q-pool' behaviour (Kroger & Klingenberg, 1973a,b (Heron et al, 1977;Ragan, 1978;. This is illustrated in Fig.…”

“…Lipid-substituted Complex I was used to investigate the effects of phospholipids on the reduction of ubiquinone analogues. It was concluded that effects on activity of phase transition of the phospholipid were exerted not directly on the protein but on the mobility and concentration of ubiquinone in the lipid phase of the system (Ragan, 1978).…”

Purification and Preliminary Characterization of Mitochondrial Complex I (NADH:Ubiquinone Reductase) from Broad Bean (Vicia faba L.)

“…The low activities measured for ubiquinone-10, -30, and -50 may be due in part to the enzyme specificity for the unknown natural acceptor in plant mitochondria and partly to the extremely low solubility of ubiquinone-H in the assay medium when n is greater than 2. This phenomenon was studied in mammalian mitochondria by Ragan (1978), who showed the importance of phospholipids in solubilizing the ubiquinone in the environment of its reduction site. In our case, preliminary experiments with phosphatidylcholine did not significantly increase the activity with ubiquinone-30, but more extensive investigations are needed, notably to determine which phospholipids should be used.…”