Assessment of the shape and molecular size of TSH-TSH receptor complexes.
Photoaffinity labelling and analysis under denaturing conditions (SDS-PAGE) have shown that the porcine TSH receptor contains an A subunit (Mr = 47,000) which forms the binding site for TSH and a B subunit (Mr = 25,000) linked to the A subunit by a disulphide bridge. In order to assess the size and shape of the receptor under non-denaturing conditions we have solubilized photoaffinity-labelled porcine TSH receptors using the small micelle-sized detergent sodium deoxycholate and analysed the preparations by sucrose density gradient centrifugation and gel filtration. Under these conditions, the cross-linked TSH-TSH receptor complex showed an S20,w of 6.4 S and a frictional ratio f/f0 of 1.8. These values were consistent with those which might be expected from an elongated protein complex with a molecular weight of about 100,000 (the value obtained by SDS-PAGE). Analysis of another thyroid membrane protein, human thyroid microsomal antigen (Mr = 110,000 by SDS-PAGE) under the same conditions gave an S20,w of 6.0 S and f/f0 = 1.3, suggesting that this protein has a compact structure. The TSH receptor A subunit cross-linked to TSH (Mr = 70,000 by SDS-PAGE) gave an S20,w of 4.6 S and f/f0 = 1.8 and these values could be compared with those obtained for the A subunit alone (S20,w = 3.6 S; f/f0 = 1.4; Mr by SDS-PAGE = 47,000) and TSH alone (S20,w = 2.6 S; f/f0 = 1.6; Mr = 28,000.(ABSTRACT TRUNCATED AT 250 WORDS)
D000345:Affinity Labels; D000818:Animals; D001386:Azides; D002417:Cattle; D002499:Centrifugation, Density Gradient; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D003432:Cross-Linking Reagents; D003840:Deoxycholic Acid; D046911:Macromolecular Substances; D008970:Molecular Weight; D011989:Receptors, Thyrotropin / Q000378:metabolism*; D013552:Swine; D013972:Thyrotropin / Q000378:metabolism*
D000345:Affinity Labels; D001386:Azides; D003432:Cross-Linking Reagents; D046911:Macromolecular Substances; D011989:Receptors, Thyrotropin; D003840:Deoxycholic Acid; C029534:hydroxysuccinimidyl-4-azidobenzoate; D013972:Thyrotropin