Functional coupling between enzymes of the chromaffin granule membrane.
The reactions of cytochrome b561 with other redox-active components of the adrenal chromaffin granule were examined using optical difference spectroscopy. It was shown that there is no direct electron transfer between the cytochrome and dopamine beta-hydroxylase, but that in the presence of ascorbate, turnover of dopamine beta-hydroxylase causes an oxidation of the cytochrome, which is partially reversed by the action of the mitochondrial NADH:A-. oxidoreductase. Thus, these three proteins may be functionally coupled via ascorbate. A quantitative study of the relationship between the redox state of the cytochrome and the ascorbate radical concentration measured by EPR showed that ascorbate reduces the cytochrome in a one-electron transfer reaction. Generation of a proton electrochemical gradient across the granule membrane causes only a small (20 mV) increase in the cytochrome midpoint potential suggesting the cytochrome is not a proton pump. The data are consistent with a model in which cytochrome b561, by reacting with ascorbate or ascorbate free radical on either side of the granule membrane, could couple the ascorbate-consuming reaction of the dopamine beta-hydroxylase inside the chromaffin granule to the ascorbate-regenerating reaction of the NADH:A-. oxidoreductase on the outer mitochondrial membrane. The H+-ATPase of the granule membrane could both drive the flow of electrons in the direction from cytosol to granule and replenish protons consumed by the turnover of dopamine beta-hydroxylase inside the granule.
D000818:Animals; D001205:Ascorbic Acid / Q000378:metabolism; D002417:Cattle; D002837:Chromaffin Granules / Q000201:enzymology*; D002838:Chromaffin System / Q000201:enzymology*; D003573:Cytochrome b Group / Q000032:analysis*; D004298:Dopamine / Q000494:pharmacology; D004299:Dopamine beta-Hydroxylase / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D004837:Epinephrine / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology; D005609:Free Radicals; D005650:Fumarates / Q000494:pharmacology; D005669:Fusaric Acid / Q000494:pharmacology; D007425:Intracellular Membranes / Q000201:enzymology*; D009243:NAD / Q000378:metabolism; D009638:Norepinephrine / Q000494:pharmacology; D010084:Oxidation-Reduction; D013053:Spectrophotometry; D014439:Tyramine / Q000494:pharmacology
D003573:Cytochrome b Group; D005609:Free Radicals; D005650:Fumarates; D009243:NAD; C023646:cytochrome b561; C032005:fumaric acid; D004299:Dopamine beta-Hydroxylase; D005669:Fusaric Acid; D005033:Ethylmaleimide; D001205:Ascorbic Acid; D004298:Dopamine; D009638:Norepinephrine; D014439:Tyramine; D004837:Epinephrine