The insertion of monoamine oxidase A into the outer membrane of rat liver mitochondria.
Human monoamine oxidase A that had been synthesized in a reticulocyte lysate translation system was capable of binding to and inserting into either rat liver mitochondria or isolated mitochondrial outer membranes. The inserted form was as resistant to proteinase K as endogenous mitochondrial monoamine oxidase A. The insertion, but not the binding, of monoamine oxidase A was prevented by depleting the reaction mixture of either ATP (with apyrase) or ubiquitin (with purified antibodies against this polypeptide). Addition of ATP or ubiquitin, respectively, to these depleted mixtures restored the insertion of the enzyme. In the absence of mitochondria, in vitro synthesized monoamine oxidase A did not catalyze its own alkylation by the mechanism-based inhibitor, [3H]clorgyline. However, both monoamine oxidase A that had been membrane-inserted in vitro and monoamine oxidase A that had been bound to the mitochondria under conditions of ATP depletion catalyzed adduct formation. Furthermore, reaction of either clorgyline or another mechanism-based inhibitor, pargyline, with the membrane-bound enzyme during ATP depletion inhibited the insertion of monoamine oxidase A when ATP was restored. These observations indicate that monoamine oxidase A acquired a catalytically active conformation on interaction with the mitochondrial outer membranes prior to its ATP and ubiquitin-dependent insertion into the membrane.
Zhuang
Z P
ZP
Department of Pharmacology, Wayne State University, Detroit, Michigan 48201.
Marks
B
B
McCauley
R B
RB
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002384:Catalysis; D003010:Clorgyline / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D019286:Endopeptidase K; D006801:Humans; D007425:Intracellular Membranes / Q000187:drug effects / Q000201:enzymology; D008930:Mitochondria, Liver / Q000187:drug effects / Q000201:enzymology*; D008995:Monoamine Oxidase / Q000235:genetics / Q000378:metabolism*; D010293:Pargyline / Q000494:pharmacology; D051381:Rats; D012697:Serine Endopeptidases / Q000378:metabolism; D014452:Ubiquitins / Q000378:metabolism
D014452:Ubiquitins; D000255:Adenosine Triphosphate; D010293:Pargyline; D008995:Monoamine Oxidase; D012697:Serine Endopeptidases; D019286:Endopeptidase K; D003010:Clorgyline