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Characterization of recombinant Saccharomyces cerevisiae telomerase core enzyme purified from yeast.从酵母中纯化的重组酿酒酵母端粒酶核心酶的特性分析。
Biochem J. 2005 Aug 15;390(Pt 1):169-76. doi: 10.1042/BJ20050208.
2
Saccharomyces cerevisiae Est3p dimerizes in vitro and dimerization contributes to efficient telomere replication in vivo.酿酒酵母Est3p在体外形成二聚体，且二聚化有助于体内端粒的有效复制。
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Saccharomyces cerevisiae telomerase subunit Est3p binds DNA and RNA and stimulates unwinding of RNA/DNA heteroduplexes.酿酒酵母端粒酶亚基Est3p可结合DNA和RNA，并刺激RNA/DNA杂合体的解旋。
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Ever shorter telomere 1 (EST1)-dependent reverse transcription by Candida telomerase in vitro: evidence in support of an activating function.念珠菌端粒酶在体外依赖于端粒酶相关蛋白1（EST1）的逆转录：支持激活功能的证据
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Est1p as a cell cycle-regulated activator of telomere-bound telomerase.Est1p作为端粒结合的端粒酶的细胞周期调控激活剂。
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The Est3 protein is a subunit of yeast telomerase.Est3蛋白是酵母端粒酶的一个亚基。
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Intracellular trafficking of yeast telomerase components.酵母端粒酶组分的细胞内运输
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Crystal structures of N-terminally truncated telomerase reverse transcriptase from fungi‡.真菌端粒酶逆转录酶 N 端截短体的晶体结构‡。
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Cdc13 is predominant over Stn1 and Ten1 in preventing chromosome end fusions.Cdc13 在防止染色体末端融合方面优于 Stn1 和 Ten1。
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Yeast telomere maintenance is globally controlled by programmed ribosomal frameshifting and the nonsense-mediated mRNA decay pathway.酵母端粒的维持受到程序性核糖体移码和无义介导的mRNA降解途径的全局控制。
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Yeast telomerase subunit Est1p has guanine quadruplex-promoting activity that is required for telomere elongation.酵母端粒酶亚基 Est1p 具有促进鸟嘌呤四链体形成的活性，该活性对于端粒延伸是必需的。
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Saccharomyces cerevisiae Est3p dimerizes in vitro and dimerization contributes to efficient telomere replication in vivo.酿酒酵母Est3p在体外形成二聚体，且二聚化有助于体内端粒的有效复制。
Nucleic Acids Res. 2006 Jan 17;34(2):407-16. doi: 10.1093/nar/gkj445. Print 2006.

本文引用的文献

1
Telomere length homeostasis is achieved via a switch between telomerase- extendible and -nonextendible states.端粒长度稳态是通过端粒酶可延长状态和不可延长状态之间的转换来实现的。
Cell. 2004 Apr 30;117(3):323-35. doi: 10.1016/s0092-8674(04)00334-4.
2
A conserved telomerase motif within the catalytic domain of telomerase reverse transcriptase is specifically required for repeat addition processivity.端粒酶逆转录酶催化结构域内一个保守的端粒酶基序是重复添加持续性所特别需要的。
Mol Cell Biol. 2003 Dec;23(23):8440-9. doi: 10.1128/MCB.23.23.8440-8449.2003.
3
Ever shorter telomere 1 (EST1)-dependent reverse transcription by Candida telomerase in vitro: evidence in support of an activating function.念珠菌端粒酶在体外依赖于端粒酶相关蛋白1（EST1）的逆转录：支持激活功能的证据
Proc Natl Acad Sci U S A. 2003 May 13;100(10):5718-23. doi: 10.1073/pnas.1036868100. Epub 2003 Apr 25.
4
N-terminal domain of yeast telomerase reverse transcriptase: recruitment of Est3p to the telomerase complex.酵母端粒酶逆转录酶的N端结构域：Est3p募集至端粒酶复合体
Mol Biol Cell. 2003 Jan;14(1):1-13. doi: 10.1091/mbc.e02-06-0327.
5
Est1p as a cell cycle-regulated activator of telomere-bound telomerase.Est1p作为端粒结合的端粒酶的细胞周期调控激活剂。
Science. 2002 Aug 9;297(5583):1023-6. doi: 10.1126/science.1074968.
6
Intracellular trafficking of yeast telomerase components.酵母端粒酶组分的细胞内运输
EMBO Rep. 2002 Jul;3(7):652-9. doi: 10.1093/embo-reports/kvf133.
7
Schizosaccharomyces pombe pfh1+ encodes an essential 5' to 3' DNA helicase that is a member of the PIF1 subfamily of DNA helicases.粟酒裂殖酵母pfh1+编码一种必需的5'至3' DNA解旋酶，它是DNA解旋酶PIF1亚家族的成员。
Mol Biol Cell. 2002 Jun;13(6):2180-91. doi: 10.1091/mbc.02-02-0021.
8
Molecular basis for telomere repeat divergence in budding yeast.芽殖酵母中端粒重复序列差异的分子基础。
Mol Cell Biol. 2001 Nov;21(21):7277-86. doi: 10.1128/MCB.21.21.7277-7286.2001.
9
Analysis of telomerase processivity: mechanistic similarity to HIV-1 reverse transcriptase and role in telomere maintenance.端粒酶持续合成能力分析：与HIV-1逆转录酶的机制相似性及其在端粒维持中的作用
Mol Cell. 2001 Jun;7(6):1201-11. doi: 10.1016/s1097-2765(01)00268-4.
10
Yeast telomerase appears to frequently copy the entire template in vivo.酵母端粒酶似乎在体内经常复制整个模板。
Nucleic Acids Res. 2001 Jun 1;29(11):2382-94. doi: 10.1093/nar/29.11.2382.

从酵母中纯化的重组酿酒酵母端粒酶核心酶的特性分析。

Characterization of recombinant Saccharomyces cerevisiae telomerase core enzyme purified from yeast.

作者信息

Liao Xin-Hua, Zhang Ming-Liang, Yang Cui-Ping, Xu Lu-Xia, Zhou Jin-Qiu

机构信息

Max-Planck Junior Research Group in the State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, 200031, China.

出版信息

Biochem J. 2005 Aug 15;390(Pt 1):169-76. doi: 10.1042/BJ20050208.

DOI:10.1042/BJ20050208
PMID:15813705
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC1184572/
Abstract

Telomerase is a cellular reverse transcriptase that elongates the single-stranded chromosome ends and oligonucleotides in vivo and in vitro. In Saccharomyces cerevisiae, Est2p (telomerase catalytic subunit) and Tlc1 (telomerase RNA template subunit) constitute the telomerase core complex. We co-overexpressed GST (glutathione S-transferase)-Est2p and Tlc1 in S. cerevisiae, and reconstituted the telomerase activity. The GST-Est2p-Tlc1 complex was partially purified by ammonium sulphate fractionation and affinity chromatography on glutathione beads, and the partially purified telomerase did not contain the other two subunits of the telomerase holoenzyme, Est1p and Est3p. The purified recombinant GST-Est2p-Tlc1 telomerase core complex could specifically add nucleotides on to the single-stranded TG(1-3) primer in a processive manner, but could not translocate to synthesize more than one telomeric repeat. The purified telomerase core complex exhibited different activities when primers were paired with the Tlc1 template at different positions. The procedure of reconstitution and purification of telomerase core enzyme that we have developed now allows for further mechanistic studies of the functions of other subunits of the telomerase holoenzyme as well as other telomerase regulation proteins.

摘要

端粒酶是一种细胞逆转录酶，可在体内和体外延长单链染色体末端和寡核苷酸。在酿酒酵母中，Est2p（端粒酶催化亚基）和Tlc1（端粒酶RNA模板亚基）构成端粒酶核心复合物。我们在酿酒酵母中共过表达了GST（谷胱甘肽S-转移酶）-Est2p和Tlc1，并重建了端粒酶活性。通过硫酸铵分级分离和在谷胱甘肽珠上的亲和色谱法对GST-Est2p-Tlc1复合物进行了部分纯化，并且部分纯化的端粒酶不包含端粒酶全酶的其他两个亚基Est1p和Est3p。纯化的重组GST-Est2p-Tlc1端粒酶核心复合物可以以连续的方式将核苷酸特异性地添加到单链TG(1-3)引物上，但不能转位以合成多个端粒重复序列。当引物与Tlc1模板在不同位置配对时，纯化的端粒酶核心复合物表现出不同的活性。我们现在开发的端粒酶核心酶的重建和纯化程序允许对端粒酶全酶的其他亚基以及其他端粒酶调节蛋白的功能进行进一步的机制研究。