Accurate DNA synthesis by Sulfolobus solfataricus DNA polymerase B1 at high temperature
Accurate DNA synthesis by Sulfolobus solfataricus DNA polymerase B1 at high temperature
复制标题
硫磺硫化叶菌 DNA 聚合酶 B1 高温下准确合成 DNA
DOI：
10.1007/s00792-009-0292-9
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发表时间：
2009
期刊：
影响因子：
2.9
通讯作者：
Huang, Li
中科院分区：
文献类型：
--
作者：
Zhang, Likui;Guo, Li;Guo, Xin;Zhan, Zhengyan;Lou, Huiqiang;Duan, Zhenhong;Huang, Li
利用基于M13的突变方法，研究了DNA聚合酶B1在生理温度附近对高温古生菌Sulfolobus solfararicus(SSO PolB1)DNA合成的准确性。SSO polB1显示出与大多数病毒、细菌和真核复制酶相似或更高的复制保真度。该酶在70℃时的保真度约为55℃时的三倍。约三分之二的酶错误是单碱基取代，其中58%是C-T转变。移码突变，主要由单碱基缺失引起，占总错误的19%。SSO polB1的一个核酸外切酶缺陷突变体的诱变性是野生型酶的三倍，这表明内在的校对功能对酶的保真度只有轻微的贡献。动力学分析表明，SSO polB1的外切酶缺陷三点突变的所有可能的错配频率在5.4×10(-5)到4.6×10(-4)之间。该酶在DNA合成中的高保真主要是基于K(M)差而不是V(Max)差。SSO polB1的这些性质与该酶在Solfararicus中作为复制酶所起的作用是一致的。
The accuracy of DNA synthesis by DNA polymerase B1 from the hyperthermophilic archaeon Sulfolobus solfataricus (Sso pol B1) at near the physiological temperature was investigated using M13-based mutational assays. Sso pol B1 showed replication fidelity similar to or higher than most viral, bacterial, and eukaryotic replicases. The fidelity of the enzyme was about three times as high at 70 degrees C as at 55 degrees C. Approximately two-thirds of the errors made by the enzyme were single-base substitutions, of which 58% were C --> T transition. Frameshift mutations, mostly resulting from single-base deletions, accounted for 19% of the total errors. An exonuclease-deficient mutant of Sso pol B1 was three times as mutagenic as the wild-type enzyme, suggesting that the intrinsic proofreading function contributed only modestly to the fidelity of the enzyme. Kinetic assays showed that the frequencies of all possible misincorporations by an exonuclease-deficient triple-point mutant of Sso pol B1 ranged from 5.4 x 10(-5) to 4.6 x 10(-4). The high fidelity of this enzyme in DNA synthesis was based primarily on K (m) difference rather than V (max) difference. These properties of Sso pol B1 are consistent with the proposed role of the enzyme as a replicase in S. solfataricus.
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DOI：
10.1074/jbc.273.36.23575
发表时间：
1998-09
期刊：
The Journal of Biological Chemistry
影响因子：
--
作者：
P. Pham;M. Olson;C. McHenry;R. Schaaper
通讯作者：
P. Pham;M. Olson;C. McHenry;R. Schaaper
影响因子：
4.4
作者：
Choi, Jeong Jin;Song, Jae-Geun;Kwon, Suk-Tae
通讯作者：
Kwon, Suk-Tae
影响因子：
2.9
作者：
DAWSON, JM;HEATLIE, PL
通讯作者：
HEATLIE, PL
影响因子：
2.8
作者：
Y. Kim;H. Lee;S. S. Bae-S.;J. Jeon;J. Lim;Yona Cho;K. Nam;S. Kang;Sang-Jin Kim;S. Kwon;Jung-Hyun Lee
通讯作者：
Y. Kim;H. Lee;S. S. Bae-S.;J. Jeon;J. Lim;Yona Cho;K. Nam;S. Kang;Sang-Jin Kim;S. Kwon;Jung-Hyun Lee
影响因子：
4.8
作者：
Lou, HQ;Duan, ZH;Huang, L
通讯作者：
Huang, L