Identification of two proteins associated with mammalian ATP synthase.
Bovine mitochondrial ATP synthase commonly is isolated as a monomeric complex that contains 16 protein subunits and the natural IF(1) inhibitor protein in substoichiometric amounts. Alternatively ATP synthase can be isolated in dimeric and higher oligomeric states using digitonin for membrane solubilization and blue native or clear native electrophoresis for separation of the native mitochondrial complexes. Using blue native electrophoresis we could identify two ATP synthase-associated membrane proteins with masses smaller than 7 kDa and isoelectric points close to 10 that previously had been removed during purification. We show that in the mitochondrial membrane both proteins are almost quantitatively bound to ATP synthase. Both proteins had been identified earlier in a different context, but their association with ATP synthase was unknown. The first one had been named 6.8-kDa mitochondrial proteolipid because it can be isolated by chloroform/methanol extraction from mitochondrial membranes. The second one had been denoted as diabetes-associated protein in insulin-sensitive tissue (DAPIT), which may provide a clue for further functional and clinical investigations.
Meyer
Björn
B
Institut für Pharmazeutische Chemie, Biozentrum, Centre of Excellence "Macromolecular Complexes," Johann Wolfgang Goethe-Universität Frankfurt, Max-von-Laue-Str. 9, D-60439 Frankfurt am Main, Germany.
Wittig
Ilka
I
Trifilieff
Elisabeth
E
Karas
Michael
M
Schägger
Hermann
H
D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D015180:Electrophoresis, Gel, Two-Dimensional; D006868:Hydrolysis; D013058:Mass Spectrometry; D008929:Mitochondria, Heart / Q000201:enzymology*; D024101:Mitochondrial Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D025261:Mitochondrial Proton-Translocating ATPases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D009943:Organophosphorus Compounds / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D015203:Reproducibility of Results; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
C043517:2-(4-isothiocyanatophenoxy)-1,3,2-dioxaphosphinene 2-oxide; D024101:Mitochondrial Proteins; D009943:Organophosphorus Compounds; D000255:Adenosine Triphosphate; D025261:Mitochondrial Proton-Translocating ATPases