Exploration of nucleotide binding sites in the mitochondrial membrane by 2-azido-[alpha-32P]ADP.
The ADP/ATP carrier of beef heart mitochondria is able to bind 2-azido-[alpha-32P]ADP in the dark with a Kd value of congruent to 8 microM. 2-Azido ADP is not transported and it inhibits ADP transport and ADP binding. Photoirradiation of beef heart mitochondria with 2-azido-[alpha-32P]ADP results mainly in photolabeling of the ADP/ATP carrier protein; photolabeling is prevented by carboxyatractyloside, a specific inhibitor of ADP/ATP transport. Upon photoirradiation of inside-out submitochondrial particles with 2-azido-[alpha-32P]ADP, both the ADP/ATP carrier and the beta subunit of the membrane-bound F1-ATPase are covalently labeled. The binding specificity of 2-azido-[alpha-32P]ADP for the beta subunit of F1-ATPase is ascertained by prevention of photolabeling of isolated F1 by preincubation with an excess of ADP.
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Boulay
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Vignais
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D000244:Adenosine Diphosphate / Q000031:analogs & derivatives* / Q000378:metabolism; D000818:Animals; D001386:Azides; D001665:Binding Sites; D002417:Cattle; D007425:Intracellular Membranes / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D008929:Mitochondria, Heart / Q000648:ultrastructure*; D000226:Mitochondrial ADP, ATP Translocases / Q000378:metabolism; D009711:Nucleotides / Q000378:metabolism*; D010777:Photochemistry; D006180:Proton-Translocating ATPases / Q000378:metabolism; D051381:Rats
D001386:Azides; D008565:Membrane Proteins; D009711:Nucleotides; D000244:Adenosine Diphosphate; C022295:2-azidoadenosine 3',5'-diphosphate; D000226:Mitochondrial ADP, ATP Translocases; D006180:Proton-Translocating ATPases