Difference in susceptibility of tyrosine residue to oxidative lodination between a thioredoxin box region and a hormonogenic region.
Peptide fragments, isolated from proteolytic cleavage of thyroglobulin at specific sites, were examined for the iodination of tyrosine residues. The 50 kDa polypeptide, which was prepared from digestion of bovine thyroglobulin and continuous preparative SDS-PAGE, was subjected to reduction with DTT and alkylation with iodoacetic acid to generate S-carboxymethylated peptide derivative, which was further hydrolysed by endoproteinase-Asp N. Peptide products were separated by RP-HPLC, and each fraction was analyzed by LC/ ESI-MS and MALDI-MS analyses. Based on the specificity of endoproteinase Asp-N and the mass spectra data, a peptide fragment turned out to correspond to a peptide, DALGCVKCPEGSYFQ (1438-1452), characterized by the presence of a thioredoxin box (CVKC) and a tyrosine residue. In addition, another peptide fragment (1453-1465) containing a thioredoxin box (CIPC) and a tyrosine residue was also observed. However, any evidence of iodination of the tyrosine residue present in these peptides was not provided. Meanwhile, tyrosine residues in the peptides, DVEEALAGKYLAGRFA (1366-1381) and DYSGLLLAFQVFLL (1290-1303) were found to be iodinated; mono- or diiodinated tyrosine residues, characteristic of a hormogenic site, existed in both peptides. In addition, the tyrosine residue in the peptide (1218-1252), corresponding to a hormonogenic site was also iodinated. Thus, there was a sharp difference of the susceptibility to oxidative iodination between the tyrosine residue in a hormonogenic site and that in a thioredoxin region. From these results, it is suggested that polypeptide region adjacent to tyrosine residues may govern the susceptibility of tyrosine to oxidative iodination.
Sok
D E
DE
College of Pharmacy, Chungnam National University, Taejon 305-764, Korea. daesok@cnu.ac.kr
Sih
C J
CJ
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D006868:Hydrolysis; D007455:Iodine / Q000737:chemistry*; D008745:Methylation; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000302:isolation & purification; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013820:Thermolysin / Q000737:chemistry; D013879:Thioredoxins / Q000737:chemistry*; D013954:Thyroglobulin / Q000737:chemistry; D013963:Thyroid Hormones / Q000378:metabolism*; D014443:Tyrosine / Q000737:chemistry*
D010446:Peptide Fragments; D013963:Thyroid Hormones; D014443:Tyrosine; D013879:Thioredoxins; D013954:Thyroglobulin; D007455:Iodine; D013820:Thermolysin