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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Protein tolerance to random amino acid change.
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Protein tolerance to random amino acid change.

机译：蛋白质对随机氨基酸变化的耐受性。

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Mutagenesis of protein-encoding sequences occurs ubiquitously; it enables evolution, accumulates during aging, and is associated with disease. Many biotechnological methods exploit random mutations to evolve novel proteins. To quantitate protein tolerance to random change, it is vital to understand the probability that a random amino acid replacement will lead to a protein's functional inactivation. We define this probability as the x factor. factors and demonstrate this method using the human DNA repair enzyme 3-methyladenine DNA glycosylase (AAG). Three gene-wide mutagenesis libraries were created, each with 10(5) diversity and averaging 2.2, 4.6, and 6.2 random amino acid changes per mutant. After determining the percentage of functional mutants in each library using high-stringency selection (>19,000-fold), the x factor was found to be 34% +/- 6%. Remarkably, reanalysis of data from studies of diverse proteins reveals similar inactivation probabilities. To delineate the nature of tolerated amino acid substitutions, we sequenced 244 surviving AAG mutants. The 920 tolerated substitutions were characterized by substitutability index and mapped onto the AAG primary, secondary, and known tertiary structures. Evolutionarily conserved residues show low substitutability indices. In AAG, beta strands are on average less substitutable than alpha helices; and surface loops that are not involved in DNA binding are the most substitutable. Our results are relevant to such diverse topics as applied molecular evolution, the rate of introduction of deleterious alleles into genomes in evolutionary history, and organisms' tolerance of mutational burden.
机译：蛋白质编码序列的诱变普遍存在。它能够进化，在衰老过程中积累，并与疾病有关。许多生物技术方法都利用随机突变来进化出新的蛋白质。要量化蛋白质对随机变化的耐受性，至关重要的是要了解随机氨基酸替代会导致蛋白质功能失活的可能性。我们将此概率定义为x因子。并使用人DNA修复酶3-甲基腺嘌呤DNA糖基化酶（AAG）证明了该方法。创建了三个全基因范围的诱变文库，每个文库具有10（5）多样性，每个突变体平均具有2.2、4.6和6.2个随机氨基酸变化。使用高严格性选择（> 19,000倍）确定每个文库中功能性突变的百分比后，发现x因子为34％+/- 6％。值得注意的是，对各种蛋白质研究数据的重新分析显示出相似的失活概率。为了描述耐受的氨基酸取代的性质，我们对244个存活的AAG突变体进行了测序。 920个耐受的取代基具有可取代性指数，并映射到AAG的一级，二级和已知三级结构上。进化上保守的残基显示出低的可取代指数。在AAG中，平均而言，β链的取代度比α螺旋低。与DNA结合不涉及的表面环是最易取代的。我们的结果与诸如应用分子进化，有害等位基因在进化史上向基因组中引入的速率以及生物体对突变负担的耐受性等多种主题有关。

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