Mechanism of Signal Transduction by Nonreceptor Protein-tyrosine Kinases

1. Src同源3域（SH 3）是由蛋白质酪氨酸激酶信号转导中蛋白质参与的集合中加入的蛋白质域。CDNA克隆编码一种新的蛋白配体到Fyn PTK的SH 3域已被孤立。克隆人被筛选小鼠胚胎cDNA库识别为带有Fyn·SH 3/GST融合蛋白的探针。新的SH 3配体蛋白质也是在成人大脑中表达的。蛋白质包含两个丰富的序列，与Fyn和SrcSH 3域、新的SH 3域和SH 2域的潜在配体序列结合在一起。对新近识别的蛋白质免疫吸收和免疫抑制的蛋白质进行抗体特异性鉴定，以及从COS-7细胞的裂解物转染经克隆的cDNA中转染的蛋白质。该蛋白质是由Fyn表达的从COS-7细胞中经经克隆的Fyn cDNA中的酪氨酸磷酸化的蛋白质。2. A70-kDa蛋白，具有高亲和力的Fyn SH 3被发现在Jurkat细胞裂解物与Fyn SH 3/GST融合蛋白的亲和力结合。蛋白质是从SDS-PAGE凝胶中提取出来的，用trypsin进行挖掘，肽是分开的。《气体相位显微平衡的结果》，即蛋白质与GAP结合磷脂蛋白密切相关，第62页。SH 3域是已知的绑定到proline-rich序列的。p62中的丰富蛋白序列被表达为GST融合蛋白，以及GST融合蛋白与各种SH 3/GST融合蛋白的结合被测试。结果表明，每个原线丰富的序列对SH 3域的一个子集都有一个特定的关联性。

1. The Src homology 3 domain (SH3) is a protein domain involved in the assembly of proteins participating in the signal transduction by protein-tyrosine kinases. A cDNA clone encoding a new protein ligand to the SH3 domain of Fyn PTK was isolated. The clone was identified by screening mouse embryo cDNA library with Fyn・SH3/GST fusion protein as a probe. The new SH3 ligand protein is also expressed in adult brain. The protein contains two proline-rich sequences with a specificity in binding to Fyn and Src SH3 domains, a new SH3 domain, and potential ligand sequences for SH2 domains. Antibody specific to the newly identified protein immunoprecipitated and immunoblotted a protein from the teratocarcinoma cell lysate and from the lysate of COS-7 cells transfected with the cloned cDNA.The protein was tyrosine-phosphorylated by Fyn expressed from cotransfected fyn cDNA in COS-7 cells. 2. A 70-kDa protein with high affinity to Fyn SH3 was found in the Jurkat cell lysate by affinity binding to Fyn SH3/GST fusion protein. The protein was eluted from SDS-PAGE gels, digested with trypsin, and the peptides were separated. The results of gas-phase microsequencing of the peptides indicated that the protein is closely related to GAP-binding phosphoprotein, p62. The SH3 domains are known to bind to proline-rich sequences. The proline-rich sequences in p62 were expressed as GST fusion proteins and the binding of the GST fusion proteins to various SH3/GST fusion proteins were examined. The results indicate that each proline-rich sequence has a specific affinity to a subset of SH3 domains.