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DNA聚合酶的催化编辑特性。

Catalytic editing properties of DNA polymerases.

作者信息

Canard B, Cardona B, Sarfati R S

机构信息

Faculté de Médecine, Unité de Recherche Associée-Centre National de la Recherche Scientifique 1462, Nice, France.

出版信息

Proc Natl Acad Sci U S A. 1995 Nov 21;92(24):10859-63. doi: 10.1073/pnas.92.24.10859.

DOI:10.1073/pnas.92.24.10859
PMID:7479898
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC40530/
Abstract

Enzymatic incorporation of 2',3'-dideoxynucleotides into DNA results in chain termination. We report that 3'-esterified 2'-deoxynucleoside 5'-triphosphates (dNTPs) are false chain-terminator substrates since DNA polymerases, including human immunodeficiency virus reverse transcriptase, can incorporate them into DNA and, subsequently, use this new 3' end to insert the next correctly paired dNTP. Likewise, a DNA substrate with a primer chemically esterified at the 3' position can be extended efficiently upon incubation with dNTPs and T7 DNA polymerase lacking 3'-to-5' exonuclease activity. This enzyme is also able to use dTTP-bearing reporter groups in the 3' position conjugated through amide or thiourea bonds and cleave them to restore a DNA chain terminated by an amino group at the 3' end. Hence, a number of DNA polymerases exhibit wide catalytic versatility at the 3' end of the nascent DNA strand. As part of the polymerization mechanism, these capabilities extend the number of enzymatic activities associated with these enzymes and also the study of interactions between DNA polymerases and nucleotide analogues.

摘要

将2',3'-双脱氧核苷酸酶促掺入DNA会导致链终止。我们报告称，3'-酯化的2'-脱氧核苷5'-三磷酸（dNTP）是假链终止底物，因为包括人类免疫缺陷病毒逆转录酶在内的DNA聚合酶可以将它们掺入DNA中，随后利用这个新的3'末端插入下一个正确配对的dNTP。同样，与缺乏3'-至-5'外切核酸酶活性的dNTP和T7 DNA聚合酶一起孵育时，3'位置经化学酯化的引物的DNA底物可以有效地延伸。这种酶还能够使用通过酰胺或硫脲键在3'位置共轭的带有dTTP的报告基团，并将它们切割以恢复由3'末端的氨基终止的DNA链。因此，许多DNA聚合酶在新生DNA链的3'末端表现出广泛的催化通用性。作为聚合机制的一部分，这些能力扩展了与这些酶相关的酶活性数量，也扩展了对DNA聚合酶与核苷酸类似物之间相互作用的研究。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e9f3/40530/a2eb87b91e45/pnas01502-0053-c.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e9f3/40530/5ac3ece0f1d4/pnas01502-0052-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e9f3/40530/3f5ed5860b02/pnas01502-0052-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e9f3/40530/b3ea8b6d6454/pnas01502-0053-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e9f3/40530/cc437fc52f78/pnas01502-0053-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e9f3/40530/a2eb87b91e45/pnas01502-0053-c.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e9f3/40530/5ac3ece0f1d4/pnas01502-0052-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e9f3/40530/3f5ed5860b02/pnas01502-0052-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e9f3/40530/b3ea8b6d6454/pnas01502-0053-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e9f3/40530/cc437fc52f78/pnas01502-0053-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e9f3/40530/a2eb87b91e45/pnas01502-0053-c.jpg

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本文引用的文献

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2.3 A crystal structure of the catalytic domain of DNA polymerase beta.2.3 DNA聚合酶β催化结构域的晶体结构。
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Crystal structure of bacteriophage T7 RNA polymerase at 3.3 A resolution.噬菌体T7 RNA聚合酶3.3埃分辨率的晶体结构。
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Enzymatic Cleavage of 3'-Esterified Nucleotides Enables a Long, Continuous DNA Synthesis.酶切 3’酯化核苷酸可实现长片段连续 DNA 合成。
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