The iodination sites of bovine thyrotropin.
Iodination of bovine thyrotropin (TSH) using a lactoperoxidase-catalyzed labeling method at pH 5.6 results in modification of both the alpha- and beta-subunits. In particular, 3 of the 5 tyrosine residues of the alpha-subunit and 9 of the 11 tyrosine residues in the beta-subunit are accessible to surface iodination. However, the reactivity of these tyrosine residues in bovine TSH toward iodination under these enzyme-catalyzed conditions follows the order alpha-Tyr-21 much greater than alpha-Tyr-92, -93, approximately equal to beta-Tyr-45, -54 greater than beta-Tyr-74 greater than beta-Tyr-18 approximately equal to beta-Tyr-112 greater than beta-Tyr-104 approximately equal to beta-Tyr-92 greater than beta-Tyr-7 greater than beta-Tyr-77. From reversed-phase high-performance liquid chromatography tryptic mapping, leucyl aminopeptidase M digestion, and microsequence analysis, it is clear that diiodination of the tyrosine residues is not favored for the beta-subunit with the exception of beta-Tyr-7, whereas diiodination was observed with alpha-Tyr-21 and alpha-Tyr-92/93. These data on iodination sites are evaluated in terms of the known receptor binding features of iodinated bovine TSH preparations as well as in terms of the surface accessibility of these specific residues as predicted from topographical algorithms based on an analysis of hydrophilic and hydrophobic regions of the subunits. The results provide an explanation for the anomalously low bound/total tracer ratio frequently observed in radioreceptor assay procedures for TSH and suggest a basis for further evaluation of the determinant loops associated with the hormone specificity of the beta-subunit.
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D006863:Hydrogen-Ion Concentration; D007455:Iodine / Q000378:metabolism*; D007553:Isotope Labeling; D007784:Lactoperoxidase / Q000378:metabolism; D046911:Macromolecular Substances; D010446:Peptide Fragments / Q000378:metabolism; D013972:Thyrotropin / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism
D046911:Macromolecular Substances; D010446:Peptide Fragments; D014443:Tyrosine; D013972:Thyrotropin; D007455:Iodine; D007784:Lactoperoxidase; D014357:Trypsin