Resolution and reconstitution of Rhodospirillum rubrum pyridine dinucleotide transhydrogenase. II. Solubilization of the membrane-bound component.
The Rhodospirillum rubrum pyridine dinucleotide transhydrogenase system is comprised of a membrane-bound component and an easily dissociable soluble factor. Active transhydrogenase complex was solubilized by extraction of chromatophores with lysolecithin. The membrane component was also extracted from membranes depleted of soluble factor. The solubilized membrane component reconstituted transhydrogenase activity upon addition of soluble factor. Various other ionic and non-ionic detergents, including Triton X-100, Lubrol WX, deoxycholate, and digitonin, were ineffectual for solubilization and/or inhibited the enzyme at higher concentrations. The solubilized membrane component was significantly less thermal stable than the membrane-bound component. None of the pyridine dinucleotide substrate affected the thermostability of the solubilized membrane-bound component, whereas NADP+ and NADPH afforded protection to membrane-bound component. NADPH stimulated trypsin inactivation of membrane-bound component to a greater extent that NADP+, but inactivation of solubilized membrane component was stimulated to the same extent by both pyridine dinucleotides. The solubilized membrane component appears to have a slightly higher affinity for soluble factor than does the membrane-bound component.
D020130:Bacterial Chromatophores / Q000201:enzymology; D008244:Lysophosphatidylcholines / Q000494:pharmacology; D009243:NAD / Q000378:metabolism; D009247:NADH, NADPH Oxidoreductases / Q000302:isolation & purification / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D012247:Rhodospirillum rubrum / Q000201:enzymology*; D012995:Solubility; D013501:Surface-Active Agents / Q000494:pharmacology; D013696:Temperature; D014357:Trypsin / Q000494:pharmacology
D008244:Lysophosphatidylcholines; D013501:Surface-Active Agents; D009243:NAD; D009249:NADP; D009247:NADH, NADPH Oxidoreductases; D014357:Trypsin