Abstract
We have proposed that phosphorylated and dephosphorylated forms of the mitochondrial sulfurtransferase, rhodanese, function as converter enzymes that interact with membrane-bound iron-sulfur centers of the electron transport chain to modulate the rate of mitochondrial respiration (Ogata, K., Dai, X., and Volini, M. (1989) J. Biol. Chem. 204, 2718-2725). In the present studies, we have explored some structural aspects of the mitochondrial rhodanese system. By sequential extraction of lysed mitochondria with phosphate buffer and phosphate buffer containing 20 mM cholate, we have shown that 30% of the rhodanese activity of bovine liver is membrane-bound. Resolution of cholate extracts on Sephadex G-100 indicates that part of the bound rhodanese is complexed with other mitochondrial proteins. Tests with the complex show that it forms iron-sulfur centers when incubated with the rhodanese sulfur-donor substrate thiosulfate, iron ions, and a reducing agent. Experiments on the rhodanese activity of rat liver mitochondria give similar results. Taken together, the findings indicate that liver rhodanese is in part bound to the mitochondrial membrane as a component of a multiprotein complex that forms iron-sulfur centers. The findings are consistent with the role we propose for rhodanese in the modulation of mitochondrial respiratory activity.
Highlights
The findings indicate that liver rhodanese is in part bound to the mitochondrial membrane as a component of a multiprotein complex that forms iron-sulfur centers
In the initial extraction steps, the lysed mitochondrial suspensions were diluted with phosphate buffer and centrifuged for removal of soluble rhodanese activity
In the later extraction steps, the mitochondrial pellets were treated with phosphate buffer containing sodium cholate to release membrane-bound rhodanese activity
Summary
We have proposed that phosphorylated and dephosphorylated forms of the mitochondrial sulfurtransferase, rhodanese, function as converter enzymes that interact with membrane-bound iron-sulfur centers of the electron transport chain to modulate the rate of mitochondrial respiration The findings indicate that liver rhodanese is in part bound to the mitochondrial membrane as a component of a multiprotein complex that forms iron-sulfur centers. Related studies have shown that the sulfur transferred by rhodanese activates the electron transport proteins, succinate dehydrogenase [13] and NADH dehydrogenase [14], as well as other enzymes of oxidative metabolism, for example xanthine oxidase [15] and NADH nitrate reductase [16]. We have shown quantitatively that in both bovine liver and rat liver mitochondria rhodanese is partitioned between membrane-bound and ! 9 .uble forms, and we have obtained evidence that part of the rhodanese is associated with other mitochondrial proteins in a high molecular weight, membrane-bound complex that forms iron-sulfur centers
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