RAF DOMAINS REQUIRED FOR TRANSFORMATION AND REGULATION OF KINASE ACTIVITY

激酶活性转化和调节所需的 RAF 结构域

• 批准号: 3916884
• 负责人: G FANNING-HEIDECKER
• 金额: --
• 依托单位: DIVISION OF CANCER EPIDEMIOLOGY AND GENETICS
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3916884
• 关键词: adenosine triphosphate; binding proteins; chemical binding; chemical structure function; complementary DNA; enzyme induction /repression; enzyme structure; gene conversion; gene expression; genetic regulation; glutamine; human tissue; laboratory mouse; lysine; molecular cloning; molecular genetics; mouse sarcoma virus; neoplasm /cancer genetics; nucleic acid sequence; oncogenes; oncogenic virus; oncoproteins; point mutation; protein engineering; protein kinase; protein sequence; site directed mutagenesis; tryptophan; viral carcinogenesis

The murine cellular homolog of the 3611-murine sarcoma virus (MSV) v-raf oncogene was isolated as cDNA clones and characterized. Comparisons at the DNA and protein levels showed that the gene is under strict selection, as only 5 of the 95 nucleotide exchanges in the 3' half of the mouse and human c-raf genes resulted in amino acid differences. Twelve nucleotide exchanges occurred during the conversion of mouse c-raf to 3611- MSV v-raf. Eight of these were in the coding sequence and resulted in four amino acid exchanges. None of the differences coincide with those observed in the activation of v-mil, the avian homolog of raf, indicating that truncation and/or protein fusion, rather than point mutations, is the major factor in oncogene activation. The importance of conserved amino acid motifs for the kinase function of the raf protein was investigated by comparing the transforming potential of mutants generated by site-directed mutagenesis. The conversion of the second lysine in the putative ATP-binding site, Gly-X-Gly-X2-Gly-X1/3-Lys Ile - Leu-Lys, to either glutamine or glutamic acid did not significantly affect the transforming efficiency of the v-raf gene, while conversion of the first lysine to tryptophan eliminated the activity.

3611-鼠肉瘤病毒的鼠细胞同源物 (MSV)将v-raf癌基因分离为cDNA克隆， 表征了 DNA和蛋白质水平的比较 结果表明，该基因是在严格的选择，因为只有5的95个 小鼠和人c-raf的3'半中的核苷酸交换 基因导致氨基酸的差异。 十二核苷酸 在小鼠c-raf向3611- MSV v-raf。 其中8个在编码序列中， 导致四个氨基酸交换。 的差异均不 与在激活v-mil中观察到的那些一致， raf同源物，表明截短和/或蛋白融合， 而不是点突变，是癌基因突变的主要因素。 activation. 保守的氨基酸基序对于 通过以下方法研究了raf蛋白的激酶功能： 比较通过以下方法产生的突变体的转化潜力： 定点诱变。 第二赖氨酸的转化， 推测的ATP结合位点Gly-X-Gly-X2-Gly-X1/3-Lys Ile - Leu-Lys对谷氨酰胺或谷氨酸的作用不显著 影响v-raf基因的转化效率， 第一个赖氨酸转化为色氨酸消除了 活动