Identification of residues in glutathione transferase capable of driving functional diversification in evolution - A novel approach to protein redesign
Identification of residues in glutathione transferase capable of driving functional diversification in evolution - A novel approach to protein redesign
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DOI：
10.1074/jbc.m211776200
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发表时间：
2003-03-07
影响因子：
4.8
通讯作者：
Mannervik, B
中科院分区：
文献类型：
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作者：
Ivarsson, Y;Mackey, AJ;Mannervik, B
蛋白质功能的进化可以通过基因复制后产生的有利的非同义密码子突变的正选择来驱动。通过观察不同相似物之间的位点特异性正向选择变化的存在和程度，可以确定导致功能变化的残基位置。我们将这一分析应用于编码Mu类谷胱甘肽转移酶的基因，它们在底物特异性上差异很大。大约3%的氨基酸残基位置，无论是靠近活性位点还是远离活性位点，都在统计学上显著的正选择变化下。相关人谷胱甘肽转移酶（GST） M1-1和GST M2-2密码子发生突变。在GST M2-2中，一个化学上保守的苏氨酸到丝氨酸的突变引起GST m1 -1特异性底物反式二苯乙烯氧化物的比活性增加1000倍，在环氧化合物替代底物苯乙烯氧化物和硝基苯基甘油的比活性增加30倍。GST M1-1的反向突变导致活性的相互降低。因此，鉴定高变密码子位置对蛋白质功能的重新设计具有强大的帮助，减少了对广泛的突变或结构知识的需求，有时还提示了可能被认为是功能保守的突变。
Evolution of protein function can be driven by positive selection of advantageous nonsynonymous codon mutations that arise following gene duplication. By observing the presence and degree of site-specific positive selection for change between divergent paralogs, residue positions responsible for functional changes can be identified. We applied this analysis to genes encoding Mu class glutathione transferases, which differ widely in substrate specificities. Approximately 3% of the amino acid residue positions, both near to and distant from the active site, are under statistically significant positive selection for change. Relevant human glutathione transferase (GST) M1-1 and GST M2-2 codons were mutated. A chemically conservative threonine to serine mutation in GST M2-2 elicited a 1,000-fold increase in specific activity with the GST M1-1-specific substrate trans-stilbene oxide and a 30-fold increase with the alternative epoxide substrates styrene oxide and nitrophenyl glycidol. The reverse mutation in GST M1-1 resulted in reciprocal decreases in activity. Thus, identification of hypervariable codon positions can be a powerful aid in the redesign of protein function, lessening the requirement for extensive mutagenesis or structural knowledge and sometimes suggesting mutations that would otherwise be considered functionally conservative.