PHOTOAFFINITY-LABELING OF LYSOSOMAL MEMBRANE-PROTEINS WITH [I-125]DIIODOTYROSINE, A SYSTEM-H LIGAND

Percoll-purified rat thyroid FRTL-5 cell lysosomes were photoaffinity- labeled with [I-125]diiodotyrosine to identify proteins which bind dii odotyrosine, a ligand for lysosomal transport system h. SDS-PAGE and a utoradiography of these membranes showed specific labeling of a 70-kDa protein and weak labeling of three smaller proteins. [I-125]Diiodotyr osine photolabeling of the 70-kDa protein was specifically competed ag ainst by ligands of lysosomal transport system h ligands. The 70-kDa p rotein was photolabeled more strongly in lysosomal membranes isolated from thyrotropin-stimulated cells when compared with those grown in th e absence of thyrotropin, consistent with previous demonstrations that thyrotropin stimulates system h transport. The 70-kDa protein may rep resent some portion of the system h carrier protein. (C) 1995 Academic Press, Inc.