Carbohydrate analysis of porcine thyroglobulin isoforms with different iodine contents

• Published in: Biochimica et biophysica acta Vol. 1336; no. 3; pp. 557 - 565
• Main Authors: Ikekita, Masahiko, Yajima, Hirofumi, Murata, Takeomi, Takatera, Kikuo, Watanabe, Tadashi, Sonoyama, Masashi, Tezuka, Yoshihiko, Ishii, Tadahiro, Tarutani, Osamu
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 20.10.1997
• Subjects: Animals; Asparagine; Asparagine-linked oligosaccharide; Carbohydrates - analysis; Chromatography, DEAE-Cellulose; Chromatography, High Pressure Liquid; Glycoprotein; Iodine - analysis; Iodoamino acid; Isoform; Mass Spectrometry; Oligosaccharides - chemistry; Oligosaccharides - isolation & purification; Sialic Acids - analysis; Swine; Thyroglobulin; Thyroglobulin - chemistry; Thyroglobulin - isolation & purification; FTIR, Fourier transform infrared; DIT, 3,5-diiodo- l-tyrosine; TPO, thyroid peroxidase; Iodoamino acid; Glycoprotein; ABOE, p-aminobenzoic acid octyl ester; Asparagine-linked oligosaccharide; ATR, attenuated total reflection; DEAE–cellulose, diethyl aminoethyl–cellulose; T4, thyroxine; TSH, thyroid stimulating hormone; HPLC, high-performance liquid chromatography; Tg, thyroglobulin; T3, 3,5,3′-triiodo- l-thyronine; Con A, concanavalin A; Thyroglobulin; SDS–PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis; Isoform; ICP/MS, inductively coupled plasma mass spectrometry; MIT, 3-monoiodo- l-tyrosine; CD, circular dichroism
• ISSN: 0304-4165; 0006-3002; 1872-8006
• DOI: 10.1016/S0304-4165(97)00070-6

To further validate the relationship between thyroid hormone formation and the carbohydrate structure of thyroglobulin (Tg), we reinvestigated the relationship between the iodine content and the asparagine-linked oligosaccharide structures of porcine Tg. Purified porcine Tg was further separated into isoforms (Tg-F1, -F2 and -F3) with a DEAE–cellulose ion-exchange chromatography column. The iodine residues, neutral sugar and sialic acid were analyzed for the separated Tg isoforms and their asparagine-linked oligosaccharide structures were analyzed. The asparagine-linked oligosaccharides were released from Tg-F1, -F2 and -F3 by hydrazinolysis and each oligosaccharide was labeled with p-aminobenzoic acid octyl ester (ABOE). The ABOE-labeled oligosaccharides from Tg-F1, -F2 and -F3 were analyzed for their relative content in oligosaccharides of each structure type by chemical methods and DEAE- and ConA high-performance liquid chromatography (HPLC) columns. As a result, it was revealed that the Tg fraction eluted at higher ionic strength from a DEAE–cellulose column is apt to contain more of each iodoamino acid, as well as total content of iodine, larger negative zeta-potential, conforming to sialic acid content in the Tg molecule and to a higher content of di-sialo-bi-antennary complex and to high mannose-type oligosaccharides. These results support the conclusion that iodine organification of the Tg molecule is correlated with asparagine-linked oligosaccharide completion.