Schnapp G, Rodi H P, Rettig W J, Schnapp A, Damm K
Department of Oncology Research, Boehringer Ingelheim Pharma KG, Birkendorfer Strasse 65,88397 Biberach an der Riss, Germany.
Nucleic Acids Res. 1998 Jul 1;26(13):3311-3. doi: 10.1093/nar/26.13.3311.
Human telomerase is a ribonucleoprotein (RNP) enzyme, comprising protein components and an RNA template that catalyses telomere elongation through the addition of TTAGGG repeats. Telomerase function has been implicated in aging and cancer cell immortalization. We report a rapid and efficient one-step purification protocol to obtain highly active telomerase from human cells. The purification is based on affinity chromatography of nuclear extracts with antisense oligonucleotides complementary to the template region of the human telomerase RNA component. Bound telomerase is eluted with a displacement oligonucleotide under mild conditions. The resulting affinity-purified telomerase is active in PCR-amplified telomerase assays. The purified telomerase complex has a molecular mass of approximately 550 kDa compared to the approximately 1000 kDa determined for the telomerase RNP in unfractionated nuclear extracts. The purification protocol provides a rapid and efficient tool for functional and structural studies of human telomerase.
人端粒酶是一种核糖核蛋白（RNP）酶，由蛋白质成分和一个RNA模板组成，该酶通过添加TTAGGG重复序列催化端粒延长。端粒酶功能与衰老和癌细胞永生化有关。我们报告了一种快速有效的一步纯化方案，可从人细胞中获得高活性端粒酶。该纯化基于用与人端粒酶RNA成分模板区域互补的反义寡核苷酸对核提取物进行亲和层析。在温和条件下用置换寡核苷酸洗脱结合的端粒酶。所得亲和纯化的端粒酶在PCR扩增的端粒酶测定中具有活性。与未分级核提取物中端粒酶RNP测定的约1000 kDa相比，纯化的端粒酶复合物分子量约为550 kDa。该纯化方案为人类端粒酶的功能和结构研究提供了一种快速有效的工具。
