The Structure of Telomerase RNA Attending Catalysis

通过补充每条染色体3'端的DNA，端粒酶是细胞永生的关键。端粒酶是一种重要的核酸聚合酶，以其RNA亚基为模板合成真核生物染色体3'端的DNA。端粒酶的产物，端粒DNA，是一个单链的高度重复的富含G的DNA，是一个专门的核蛋白复合物称为端粒的一部分。端粒的维持是必不可少的，因为端粒保护染色体末端免受DNA修复机制的错误识别和随后的溶核降解。端粒酶催化机制的一些特征仍然是神秘的，在这个项目中进行的研究应该阐明端粒酶功能的结构基础。具体而言，使用四膜虫嗜热端粒酶RNA（tTR）作为模型，研究人员将阐明端粒酶RNA的作用，通过使用各种高分辨率的生化技术。在第一个具体目标中，将使用通过引物延伸（SHAPE）分析的选择性2 '-羟基酰化在溶液和催化活性络合物中研究tTR的结构。在第二个目标中，将使用核苷酸类似物干扰作图来鉴定整个tTR中的重要官能团，这是一种高通量方法，允许同时询问RNA的每个核苷酸的官能团。在最终目标中，根据目标1和2的数据设计的tTR位点特异性突变体将用于证实这些发现。这项研究将由学生进行，包括科学界代表性不足的群体的成员。此外，参加少数民族暑期研究项目的学生也将参加该项目。该项目的资金将为学生提供用品和研究津贴。 此外，端粒酶是一种重要的酶，影响许多生物现象，包括衰老，因此对它的了解可以在许多方面造福社会。

By replenishing the DNA at the 3' end of each chromosome, the enzyme telomerase is one key to cellular immortality. Functionally, telomerase is a remarkable nucleic acid polymerase that uses its integral RNA subunit as the template for the synthesis of the DNA located at the 3' end of eukaryotic chromosomes. The product of telomerase, telomeric DNA, is a single strand of highly repetitive G-rich DNA that is part of the specialized nucleoprotein complex called the telomere. Telomere maintenance is essential because the telomere guards the chromosome ends from mistaken recognition by the DNA repair machinery and consequent nucleolytic degradation. Several features of the catalytic mechanism of telomerase remain mysterious, and the research conducted in this project should illuminate the structural basis for telomerase function. Specifically, using Tetrahymena thermophila telomerase RNA (tTR) as a model, the investigator will elucidate the role of telomerase RNA through the use of a variety of high resolution biochemical techniques. In the first specific aim the structure of tTR will be studied both in solution and in the catalytically active complex using selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE). In the second aim important functional groups throughout tTR will be identified using nucleotide analog interference mapping, a high throughput approach that allows simultaneous interrogation of functional groups of each nucleotide of an RNA. In the final aim, site-specific mutants of tTR, designed based on data from aims 1 and 2, will be used to corroborate those findings.This research will be conducted by students, including members of groups underrepresented in science. Furthermore, students who participate in a minority summer research program are expected to also participate in this project. Funding for the project will provide students with supplies and research stipends. Furthermore, telomerase is an important enzyme that affects many biological phenomena, including that of aging, and thus its understanding can benefit society in many ways.