Myoglobin and mitochondria: oxymyoglobin interacts with mitochondrial membrane during deoxygenation.

The rates of oxygen uptake by rat liver mitochondria (MC) (native coupled, freshly frozen, and uncoupled by FCCP) have been measured polarographically in the absence (V(0)) or presence (V(1)) of 0.11-0.25 mM sperm whale MbO2. Under the same standard conditions, the rate of sperm whale MbO2 deoxygenation (V(2)) has been studied spectrophotometrically in the presence of respiring MC. For freshly frozen MC, the dependence of V(1) and V(2) on the overall charge of MbO2 has been investigated at pH 5.6-7.6, and the influence of other differently charged proteins (apomyoglobin, egg lysozyme, lactalbumin, and BSA) has been studied at pH 7.4. It is shown that the rate of mitochondrial respiration in the presence of MbO2 increases by 10-30% (V(1) > V(0)). No myoglobin effect is observed for FCCP-uncoupled MC (V(max) does not change). The rate of MbO2 deoxygenation is equal to the rate of oxygen uptake by mitochondria (V(2)/V(1) ~ 1 at pH 7.2-7.5). At varying pH < 7.2, the V(2) values become markedly higher than V(1), evidently due to the increased MbO2 positive charge and its stronger interaction with negatively charged mitochondrial membrane. At pH 7.4, on the contrary, V(2) is twice lower than V(1) in the case of negatively charged CM-MbO2 (pI 5.2), which has carboxymethylated histidines. Positively charged lysozyme (pI 11) strongly inhibits MbO2 deoxygenation (V(2)) without affecting oxygen uptake by MC (V(0) and V(1)). At the same time, apomyoglobin (pI 8.5), which is structurally very similar to the holoprotein, and both negatively charged lactalbumin (pI 4.4) and BSA (pI 4.7) have no substantial influence on V(2) and V(1). The MC membrane evidently has no specific sites for the interaction with myoglobin. Rather, the protein contacts with phospholipids of the outer membrane during MbO2 deoxygenation, and electrostatic interactions are of great importance for this process.