Nemoto N, Kodama K, Tazawa A, Masahito P, Ishikawa T
Differentiation. 1986;32(1):17-23. doi: 10.1111/j.1432-0436.1986.tb00551.x.
We cloned ras-related sequences from goldfish genomic libraries constructed as recombinants using the lambda phage. Restriction enzyme mapping of the clones obtained revealed three kinds of ras-related sequences among approximately 350,000 genomic clones. One of these clones was partially sequenced. Comparison with the nucleotide sequences of mammalian ras genes showed that the determined sequences covered the predicted amino acid coding regions and parts of the intervening regions. The predicted amino acid sequences of the cloned ras-related goldfish gene suggested that the coding region is localized separately in DNA, and that its exon-intron boundaries are exactly the same as those of corresponding mammalian genes. The nucleotide and amino acid sequences of the goldfish ras-related gene may have extensive homologies to mammalian p 21 protein. Among the three mammalian ras proteins, the predicted amino acid sequence of the sequenced ras-related goldfish clone is most closely homologous (96%) to the Kirsten ras protein. Differences in the predicted amino acid sequence were greatest in the sequence predicted from the fourth exon; fewer differences were found in the sequence from the third exon, and only slight or no differences were found in the sequence predicted for the first and second exons. The 12th and 61st amino acids from the N-terminal of the protein, which are thought to be critical positions for GTP binding and catalysis, are both conserved in the goldfish protein.(ABSTRACT TRUNCATED AT 250 WORDS)
我们从以λ噬菌体构建的重组金鱼基因组文库中克隆了与ras相关的序列。对所获得克隆的限制性酶切图谱分析显示,在大约350,000个基因组克隆中存在三种与ras相关的序列。对其中一个克隆进行了部分测序。与哺乳动物ras基因的核苷酸序列比较表明,所测定的序列覆盖了预测的氨基酸编码区及部分间隔区。克隆的金鱼ras相关基因的预测氨基酸序列表明,编码区在DNA中是分开定位的,并且其外显子-内含子边界与相应哺乳动物基因的完全相同。金鱼ras相关基因的核苷酸和氨基酸序列可能与哺乳动物p21蛋白有广泛的同源性。在三种哺乳动物ras蛋白中,已测序的金鱼ras相关克隆的预测氨基酸序列与Kirsten ras蛋白的同源性最高(96%)。预测氨基酸序列的差异在从第四外显子预测的序列中最大;在从第三外显子预测的序列中差异较少,而在为第一和第二外显子预测的序列中仅发现轻微差异或无差异。该蛋白N端的第12和61个氨基酸,被认为是GTP结合和催化的关键位置,在金鱼蛋白中均保守。(摘要截短于250词)