A long-chain fatty acyl CoA photolabel, 2-azido [P-32]palmitoyl CoA, w
as synthesized and its covalent interaction with mitochondrial membran
e proteins examined, On binding of 2-azido [P-32]palmitoyl CoA to beef
heart mitochondria, two polypeptides were primarily labeled, the 30 k
Da ADP/ATP carrier and a 41 kDa protein of unknown identity, Carboxyat
ractyloside and palmitoyl CoA completely protected against labeling of
the 30 kDa protein indicating that it was the ADP/ATP carrier. With i
nverted submitochondrial particles, only the 30 kDa polypeptide was la
beled by 2-azido [P-32]palmitoyl CoA. The labeling was inhibited by bo
ngkrekic acid and palmitoyl CoA but not carboxyatractyloside, providin
g evidence that the ADP/ATP carrier was covalently bound from the matr
ix side of the membrane, In brown adipose tissue mitochondria, 2-azido
[P-32]palmitoyl CoA photolabeled the ADP/ATP carrier and the 32 kDa u
ncoupling protein with some minor labeling of 36 and 68 kDa polypeptid
es, The results indicated that this physiological photolabeling reagen
t with the azido group on the CoA portion of the molecule interacts li
ke 2-azido ADP with nucleotide binding sites of a number of important
enzymes in cell metabolism, Moreover, the evidence strongly supports t
he hypothesis that long chain fatty acyl CoA esters are natural ligand
s for key nucleotide binding proteins.