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T7 DNA聚合酶-核酸外切酶对受损DNA的消化作用。

Digestion of damaged DNA by the T7 DNA polymerase-exonuclease.

作者信息

Koehler D R, Hanawalt P C

机构信息

Department of Biological Sciences, Stanford University, CA 94305-5020.

出版信息

Biochem J. 1993 Jul 15;293 ( Pt 2)(Pt 2):451-3. doi: 10.1042/bj2930451.

DOI:10.1042/bj2930451
PMID:8343124
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC1134381/
Abstract

We have investigated the 3'-5'-exonuclease activity of phage T7 DNA polymerase for its usefulness as an approach for the detection of lesions in DNA. Unlike the T4 DNA polymerase-exonuclease, which is commonly used to map the position and frequency of lesions in very small DNA fragments, T7 DNA polymerase-exonuclease is able to hydrolyse almost completely the large fragments from KpnI-restricted mammalian DNA. However, we found that the exonuclease was also able to hydrolyse DNA containing several kinds of lesions: cyclobutane pyrimidine dimers, thymine glycols, and mono-adducts of 4'-hydroxymethyl-4,5',8-trimethylpsoralen and 5'-methyl-isopsoralen. Modifications of the reaction conditions did not significantly alter the extent of hydrolysis. These properties distinguish the T7 DNA polymerase-exonuclease from the T4 DNA polymerase-exonuclease and make the T7 DNA polymerase-exonuclease unsuitable for detecting several types of lesions in DNA.

摘要

我们研究了噬菌体T7 DNA聚合酶的3'-5'-外切核酸酶活性，以评估其作为检测DNA损伤方法的实用性。与常用于绘制非常小的DNA片段中损伤位置和频率的T4 DNA聚合酶-外切核酸酶不同，T7 DNA聚合酶-外切核酸酶能够几乎完全水解来自KpnI酶切的哺乳动物DNA的大片段。然而，我们发现该外切核酸酶也能够水解含有多种损伤的DNA：环丁烷嘧啶二聚体、胸腺嘧啶乙二醇，以及4'-羟甲基-4,5',8-三甲基补骨脂素和5'-甲基异补骨脂素的单加合物。反应条件的改变并没有显著改变水解程度。这些特性将T7 DNA聚合酶-外切核酸酶与T4 DNA聚合酶-外切核酸酶区分开来，使得T7 DNA聚合酶-外切核酸酶不适用于检测DNA中的几种损伤类型。

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