Doi N, Itaya M, Yomo T, Tokura S, Yanagawa H
Graduate School of Environmental Earth Sciences, Hokkaido University, Sapporo, Japan.
FEBS Lett. 1997 Feb 3;402(2-3):177-80. doi: 10.1016/s0014-5793(96)01522-0.
Random sequences of 120-130 amino acid residues were inserted into a surface loop region of Escherichia coli RNase HI. This library was screened and about 10% of the clones were found to retain RNase H activity. Subsequent random mutagenesis led to an increase in RNase H activity and solubility of the protein. The inserted regions were found not to contribute to the secondary structure of the mutant protein. The high frequency of insertion of flexible sequences and the increase in the protein's function by further mutagenesis simulate one of the events in protein evolution.
将120 - 130个氨基酸残基的随机序列插入大肠杆菌核糖核酸酶H I的一个表面环区域。对该文库进行筛选，发现约10%的克隆保留核糖核酸酶H活性。随后的随机诱变导致核糖核酸酶H活性增加以及蛋白质溶解性增强。发现插入区域对突变蛋白的二级结构没有贡献。柔性序列的高频率插入以及通过进一步诱变使蛋白质功能增强，模拟了蛋白质进化中的一个事件。
