单链结合蛋白可提高体外DNA合成的保真度。
Single-strand binding protein enhances fidelity of DNA synthesis in vitro.
作者信息
Kunkel T A, Meyer R R, Loeb L A
出版信息
Proc Natl Acad Sci U S A. 1979 Dec;76(12):6331-5. doi: 10.1073/pnas.76.12.6331.
Abstract
The effect of Escherichia coli single-strand binding protein on the accuracy of in vitro DNA synthesis has been determined by using two independent methods. By using the synthetic polynucleotide poly[d(A-T)] and measuring dGTP misincorporation or by using phi X174 DNA and measuring nucleotide substitutions, we found that binding protein increases the fidelity of DNA synthesis by as much as 10-fold. This increase is observed with DNA polymerases of divergent sources and is progressive with increasing concentration of binding protein. The increased accuracy observed with DNA polymerases lacking a 3' leads to 5' exonuclease points to a mechanism other than augmented proofreading. In accord with the properties of single-strand binding proteins, it is suggested that increased fidelity is a result of enhanced base selection by the DNA polymerase, resulting from increased rigidity of the template due to its interaction with binding protein.
摘要
通过两种独立的方法测定了大肠杆菌单链结合蛋白对体外DNA合成准确性的影响。使用合成多核苷酸聚[d(A-T)]并测量dGTP错掺入，或者使用φX174 DNA并测量核苷酸取代，我们发现结合蛋白可将DNA合成的保真度提高多达10倍。在来源不同的DNA聚合酶中均观察到这种增加，并且随着结合蛋白浓度的增加而逐渐增加。在缺乏3'至5'核酸外切酶的DNA聚合酶中观察到的准确性提高指向了一种不同于增强校对的机制。与单链结合蛋白的特性一致，有人提出保真度的提高是由于DNA聚合酶增强了碱基选择的结果，这是由于模板与结合蛋白相互作用导致模板刚性增加所致。
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