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Functional analysis of domain structures of human hepatocyte growth factor

人肝细胞生长因子结构域的功能分析

基本信息

批准号：
02454540
负责人：
KITAMURA Naomi
金额：
$ 4.35万
依托单位：
Kansai Medical University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1990
资助国家：
日本
起止时间：
1990 至 1991
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-02454540/
关键词：
Hepatocyte growth factor Domain structures C-met protein クリングル構造

项目摘要

Human hepatocyte growth factor (hHGF) is a multi-functional protein. hHGF consists of characteristic structural domains ; it has four kringle domains in the heavy chain and a serine proteaselike domain in the light chain. To elucidate the role of these domain structures, we prepared mutant proteins lacking each of these domains and examined their biological activities for stimulation of hepatocyte DNA synthesis, inhibition of MethA cell growth and induction of MDCK cell dissociation. We also examined their interactions with the c-met/HGF receptor by displacement analysis and by analysis of levels of tyrosine phosphorylation. The mutant proteins lacking the Nterminal, the first kringle or the second kringle domain were not biologically effective and could not displace hHGF bound to the c-met/HGF receptor. The results indicate that these domains are necessary for the biological activities of hHGF mediated by binding to the c-met/HGF receptor. The mutant proteins lacking the third or fourth kringle domain moderately retained biological activities and the receptor binding. The relative levels of the tyrosine phosphorylation of the c-met/HGF receptor by these mutant proteins correlated well with the relative potencies of the biological activities when compared with the wild-type hHGF. The mutant protein lacking the light chain was not effective in the biological activities and tyrosine phosphorylation of the c-met/HGF receptor, but displaced hHGF bound to the c-met/HGF receptor. These results suggest that the heavy chain plays an important role in the interaction of hHGF with the c-met/HGF receptor and that the light chain is further required for the tyrosine phosphorylation of the c-met/HGF receptor.

人肝细胞生长因子（hHGF）是一种多功能蛋白质。 hHGF 由特征结构域组成；它在重链中有四个三环结构域，在轻链中有一个丝氨酸蛋白酶样结构域。为了阐明这些结构域结构的作用，我们制备了缺乏每个结构域的突变蛋白，并检查了它们刺激肝细胞 DNA 合成、抑制 MethA 细胞生长和诱导 MDCK 细胞解离的生物活性。我们还通过置换分析和酪氨酸磷酸化水平分析检查了它们与 c-met/HGF 受体的相互作用。缺乏N末端、第一三环结构域或第二三环结构域的突变蛋白没有生物学活性并且不能取代与c-met/HGF受体结合的hHGF。结果表明，这些结构域对于 hHGF 通过与 c-met/HGF 受体结合介导的生物活性是必需的。缺乏第三或第四三环结构域的突变蛋白适度保留了生物活性和受体结合。与野生型hHGF相比，这些突变蛋白对c-met/HGF受体的酪氨酸磷酸化的相对水平与生物活性的相对效力密切相关。缺乏轻链的突变蛋白对c-met/HGF受体的生物活性和酪氨酸磷酸化无效，但取代了与c-met/HGF受体结合的hHGF。这些结果表明重链在hHGF与c-met/HGF受体的相互作用中起重要作用，并且轻链是c-met/HGF受体的酪氨酸磷酸化进一步需要的。