Marriq C, Lejeune P J, Venot N, Vinet L
Unité 38, Institut National de la Santé et de la Recherche Médicale, Faculté de Médecine, Marseilles, France.
Mol Cell Endocrinol. 1991 Oct;81(1-3):155-64. doi: 10.1016/0303-7207(91)90214-d.
The 22 kDa fragment (Asn1-Met171) purified from iodine-poor human thyroglobulin (hTg) is capable by itself to synthesize thyroxine at Tyr5, the preferential hormonogenic acceptor site of the protein, after iodination in vitro. To identify the corresponding donor site in this model we studied the fate of the six Tyr residues present in the 22 kDa peptide after in vitro hormone synthesis. Structural studies of the tyrosyl peptides showed that Tyr5 was the only thyroxine-forming site, the other tyrosines (29, 89, 97 and 107) were noniodinated and Tyr130 was recovered in alanine form after CNBH4 treatment of the Tyr130-containing peptide. Taking into account that alanine could arise from aminoreduced pyruvate species, these results showed that in the 22 kDa fragment (1) hormone formation involves the couple Tyr5 (acceptor)-Tyr130 (donor), and (2) dehydroalanine, the resultant product of donor tyrosine after hormone synthesis, has evolved in pyruvoyl form. To test whether Tyr130 could also act as donor in hTg hormone synthesis, the 22 kDa peptide was isolated from hTg iodinated under conditions leading to iodotyrosine formation followed or not by hormone formation and the tyrosyl peptides were analyzed. After hTg iodination and before coupling (i.e. hormone synthesis) only Tyr5 and Tyr130 were recovered in iodotyrosine form; after coupling thyroxine was found at Tyr5 whereas Tyr130 disappeared. Taken together these results, correlated with the previously reported cleavage of hTg chain at Tyr130 occurring during in vivo hormone synthesis, support the theory that the couple Tyr5 (acceptor)-Tyr130 (donor) would be the preferential hormonogenic site in human Tg.
从缺碘人甲状腺球蛋白（hTg）中纯化得到的22 kDa片段（Asn1 - Met171），在体外碘化后，其自身能够在该蛋白的优先激素生成受体位点Tyr5处合成甲状腺素。为了在该模型中确定相应的供体位点，我们研究了体外激素合成后22 kDa肽段中六个Tyr残基的去向。酪氨酰肽段的结构研究表明，Tyr5是唯一形成甲状腺素的位点，其他酪氨酸（29、89、97和107）未碘化，并且在对含Tyr130的肽段进行CNBH4处理后，Tyr130以丙氨酸形式回收。考虑到丙氨酸可能源自氨基还原的丙酮酸物种，这些结果表明，在22 kDa片段中：（1）激素形成涉及Tyr5（受体）-Tyr130（供体）这一对；（2）激素合成后供体酪氨酸的产物脱氢丙氨酸已演变成丙酮酰形式。为了测试Tyr130在hTg激素合成中是否也能作为供体，从在导致碘酪氨酸形成随后或不随后形成激素的条件下碘化的hTg中分离出22 kDa肽段，并对酪氨酰肽段进行分析。在hTg碘化后且在偶联（即激素合成）之前，仅Tyr5和Tyr130以碘酪氨酸形式回收；偶联后，在Tyr5处发现甲状腺素，而Tyr130消失。这些结果与先前报道的体内激素合成过程中hTg链在Tyr130处的裂解相关，支持了Tyr5（受体）-Tyr130（供体）这一对是人类Tg中优先激素生成位点的理论。
