Euplotes telomerase:: Evidence for limited base-pairing during primer elongation and dGTP as an effector of translocation
Euplotes telomerase:: Evidence for limited base-pairing during primer elongation and dGTP as an effector of translocation
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DOI：
10.1021/bi972988o
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发表时间：
1998-04-14
期刊：
影响因子：
2.9
通讯作者：
Cech, TR
中科院分区：
文献类型：
--
作者：
Hammond, PW;Cech, TR
真核生物染色体末端的端粒重复序列由核糖核蛋白酶端粒酶维持。端粒DNA引物在活性位点（包括与RNA模板的碱基配对）和第二个锚位点处被端粒酶结合。通过测量它们的解离速率（k（off）），使用涉及在锚位点处的光交联的测定来确定羽状游仆虫引物-端粒酶复合物的稳定性。改变引物长度，并以系统的方式引入错配取代。我们观察到k（off）不与引物长度成比例，如对于累积的引物-模板碱基配对所预期的。这表明端粒酶维持着或多或少的恒定数量的碱基对，类似于RNA聚合酶维持的转录泡。通过比较引物-端粒酶复合物的实验k（off）与模型DNA-RNA双链体的实验k（off）来估计上限。所有的结合能都可以归因于10或11个碱基对;或者，可以存在
The telomeric sequence repeats at the ends of eukaryotic chromosomes are maintained by the ribonucleoprotein enzyme telomerase. Telomeric DNA primers are bound by telomerase both at the active site, which includes base-pairing with the RNA template, and at a second anchor site. The stabilities of Euplotes aediculatus primer-telomerase complexes were determined by measuring their dissociation rates (k(off)), using an assay involving photo-cross-linking at the anchor site. The primer length was varied, and mismatched substitutions were introduced in a systematic manner. We observed that k(off) does not scale with primer length as expected for accumulated primer-template base-pairing. This suggests that telomerase maintains a more-or-less constant number of base pairs, similar to the transcription bubble maintained by RNA polymerase. An upper limit was estimated by comparing the experimental k(off) for the primer-telomerase complex to that of a model DNA-RNA duplex. All the binding energy could be attributed to 10 or 11 base pairs; alternatively, there could be