Nucleotide modification at the gamma-phosphate leads to the improved fidelity of HIV-1 reverse transcriptase.
Nucleotide modification at the gamma-phosphate leads to the improved fidelity of HIV-1 reverse transcriptase.
复制标题
γ-磷酸盐的核苷酸修饰可改善HIV-1逆转录酶的保真度。
DOI：
10.1093/nar/gki779
复制
发表时间：
2005
影响因子：
14.9
通讯作者：
Hardin, SH
中科院分区：
文献类型：
--
作者：
Mulder, BA;Anaya, S;Yu, PL;Lee, KW;Nguyen, A;Murphy, J;Willson, R;Briggs, JM;Gao, XL;Hardin, SH
HIV-1逆转录酶(HIV-RT)区分正确和错误核苷酸的机制尚不清楚。合成了含有1-氨基-5-磺酸(ANS)的化学修饰核苷酸，并将其连接到γ-磷酸上，用于探测这种容易出错的聚合酶对核苷酸的选择。引物延伸反应提供了聚合酶能够结合伽马修饰的核苷酸的直接证据。正向突变分析显示，经修饰的核苷酸的突变频率降低了6倍，特定的碱基替换大大减少或消除。分子模拟说明了聚合酶活性部位内的关键残基与修饰核苷酸之间的潜在相互作用。我们的数据表明，使用修饰的核苷酸可以提高逆转录酶的保真度，我们建议对γ-磷酸的特定修饰可能有助于设计新的抗病毒疗法，或者更广泛地，作为一种工具来定义聚合酶活性部位对核苷酸选择性的结构作用。
The mechanism by which HIV-1 reverse transcriptase (HIV-RT) discriminates between the correct and incorrect nucleotide is not clearly understood. Chemically modified nucleotides containing 1-aminonaphthalene-5-sulfonate (ANS) attached to their γ-phosphate were synthesized and used to probe nucleotide selection by this error prone polymerase. Primer extension reactions provide direct evidence that the polymerase is able to incorporate the gamma-modified nucleotides. Forward mutation assays reveal a 6-fold reduction in the mutational frequency with the modified nucleotides, and specific base substitutions are dramatically reduced or eliminated. Molecular modeling illustrates potential interactions between critical residues within the polymerase active site and the modified nucleotides. Our data demonstrate that the fidelity of reverse transcriptase is improved using modified nucleotides, and we suggest that specific modifications to the γ-phosphate may be useful in designing new antiviral therapeutics or, more generally, as a tool for defining the structural role that the polymerase active site has on nucleotide selectivity.
登录
查看更多内容
影响因子：
56.9
作者：
COFFIN, JM
通讯作者：
COFFIN, JM
影响因子：
4.8
作者：
Fisher, TS;Prasad, VR
通讯作者：
Prasad, VR
DOI：
10.1073/pnas.92.14.6329
发表时间：
1995-07-03
影响因子：
11.1
作者：
HORLACHER, J;HOTTIGER, M;BENNER, SA
通讯作者：
BENNER, SA
影响因子：
4.8
作者：
Arzumanov, AA;Semizarov, DG;Krayevsky, AA
通讯作者：
Krayevsky, AA
影响因子：
4.8
作者：
Harris, D;Kaushik, N;Pandey, VN
通讯作者：
Pandey, VN