Photoaffinity labeling of hamster brown adipose tissue mitochondria by an [125I] coenzyme A derivative: differential interaction with the uncoupling protein and ADP/ATP carrier

Abstract

We have recently synthesized an azido [125I] CoA photolabel, N-(3-iodo-4-azidophenyl propionamide) cysteinyl-5-(2'thiopyridyl cysteine) CoA that specifically labeled the ADP/ATP carrier in beef heart mitochondria. In this study brown adipose tissue mitochondria were photolabeled with the azido [125I] ACT-CoA derivative with or without inhibitors. SDS gel electrophoresis and autoradiography of the separated proteins revealed exclusive photolabeling of two polypeptides corresponding to the ADP/ATP carrier and uncoupling protein. In the presence of carboxyatracytloside only the 32 kD UCP was labeled by [125I] ACT-CoA, whereas preincubation with GDP resulted in exclusive photolabeling of the 30 kD ADP/ATP carrier. Palmitoyl CoA but not palmitic acid inhibited photolabeling of both polypeptides.