STRUCTURAL STUDIES OF TELOMERIC PROTEIN--SSDNA COMPLEXES

端粒蛋白--SSDNA复合物的结构研究

• 批准号: 6164308
• 负责人: STEVE C SCHULTZ
• 金额: $ 23.11万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-05-01 至 2004-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6164308
• 关键词: DNA; DNA binding protein; X ray crystallography; chemical kinetics; crystallization; intermolecular interaction; molecular assembly /self assembly; nucleic acid structure; nucleoproteins; protein binding; protein protein interaction; protein structure function; structural biology; telomere

Telomeres are specialized protein-DNA structures that compose the very ends of linear eukaryotic chromosomes. Telomeres are essential for chromosome stability in that they protect the ends of chromosomes from degradation and recombination and participate in processes that restore terminal nucleotides that are lost during DNA replication. When telomere length and structure are not properly maintained, cells lose their ability to divide and become senescent. Lack of telomere maintenance in normal human cells appears to limit their proliferative potential, which has important implications for the processes of aging and cancer. We are investigating the precise three-dimensional structures of single-strand DNA-protein complexes that form the extreme 3'-termini of telomeres and, therefore, constitute the very ends of chromosomes. The ends of Oxytricha nova telomeres contain a 16 nucleotide single- strand 3'-DNA extension of the telomeric (TTTTGGGG)n DNA that is bound tightly by a two subunit protein referred to as the telomere binding protein. We have determined crystal structures of a ternary complex containing the alpha and beta subunits of the O. nova telomere binding protein complexed with telomeric GGGGTTTTGGGG DNA (the alpha-beta-DNA complex) and of the alpha subunit complexed with TTTTGGGG DNA (the alpha-DNA complex). We now propose to: (i) complete our structural work on these unique protein-ssDNA complexes (ii) investigate biochemically and structurally the many interesting protein-protein and protein- nucleic acid interactions observed in these complexes, and (iii) explore the intriguing possibility the the alpha-beta-DNA ternary complex forms via an ordered assembly pathway as suggested by biochemical experiments. These studies will yield new insights into telomere structure and function, single-strand DNA binding and recognition, and kinetic pathways for assembling complex macromolecular nucleoprotein structures.

端粒是一种特殊的蛋白质-DNA结构，组成线性真核生物染色体的末端。 端粒对染色体稳定性至关重要，因为它们保护染色体末端免于降解和重组，并参与恢复DNA复制期间丢失的末端核苷酸的过程。 当端粒的长度和结构没有得到适当的维持时，细胞就失去了分裂和衰老的能力。 正常人类细胞中缺乏端粒维持似乎限制了它们的增殖潜力，这对衰老和癌症的过程具有重要意义。 我们正在研究单链DNA-蛋白质复合物的精确三维结构，这些复合物形成端粒的末端，因此构成染色体的末端。新尖孢霉端粒的末端含有端粒（TTTTGGGG）n DNA的16个核苷酸的单链3 '-DNA延伸，其被称为端粒结合蛋白的两个亚基蛋白紧密结合。 我们已经确定了含有O的α和β亚基的三元复合物的晶体结构。nova端粒结合蛋白与端粒GGGGTTTTGGGG DNA复合（α-β-DNA复合物）和α亚基与TTTTGGGG DNA复合（α-DNA复合物）。 我们现建议：（i）完成我们在这些独特的蛋白质-ssDNA复合物上的结构工作;（ii）从生物化学和结构上研究在这些复合物中观察到的许多有趣的蛋白质-蛋白质和蛋白质-核酸相互作用;（iii）探索有趣的可能性，如生物化学实验所建议的，α-β-DNA三元复合物通过有序组装途径形成。这些研究将产生新的见解端粒的结构和功能，单链DNA的结合和识别，组装复杂的大分子核蛋白结构的动力学途径。