SPECTROSCOPY OF HEAVY ATOM-PERTURBED BIOPOLYMERS

重原子扰动生物聚合物的光谱学

• 批准号: 3249970
• 负责人: AUGUST H MAKI
• 金额: $ 12.52万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 1981
• 资助国家: 美国
• 起止时间: 1981-01-01 至 1998-08-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3249970
• 关键词: DNA binding protein; antibiotics; chemical binding; electron spin resonance spectroscopy; fluorescent dye /probe; heavy metals; human immunodeficiency virus 1; intermolecular interaction; isomorphous substitution; linear energy transfer; methyltransferase; mutagens; nucleic acid sequence; nucleocapsid; nucleoproteins; nucleotide analog; phosphorescence; protein structure function; simian immunodeficiency virus; structural biology; thermodynamics; transcription factor; triplet state; tryptophan; virus protein

The broad long term objectives of this research project are to obtain information at the molecular level about protein-nucleic acid interaction. Of particular importance is the furthering of our understanding of factors influencing nucleic acid sequence-selectivity, and the stability of specific complexes. The interaction between proteins and nucleic acids is a fundamental component of cellular processes, as well as being essential for the assembly and functioning of infectious agents such as mammalian retroviruses. The major specific aim of the proposed research is the study of nucleic acid complexes of the nucleocapsid proteins of type 1 human immunodeficiency virus (an infectious agent leading to AIDS), HIV-1 NC p7, and of simian immunodeficiency virus, SIV NC p8. Along with all known retroviral NC proteins, p7 and p8 contain zinc finger CCHC arrays that include highly conserved aromatic residues such as trp and phe. These Zn finger arrays have been implicated in specific nucleic acid binding and packaging. Our research will focus on the use of optical detection of triplet state magnetic resonance (ODMR) of p7 and p8 complexes with heavy atom-derivatized nucleic acids to determine whether aromatic stacking interactions are involved in binding. The triplet state zero-field splitting shifts induced in trp by complex formation with a variety of nucleic acids will be used to quantitatively evaluate the contribution of aromatic stacking interactions to complex stability, and thereby, their influence on sequence selectivity. This goal will require further investigations of echinomycin (a DNA bis-intercalating antibiotic) analogs by ODMR spectroscopy as models of stacking-induced sequence selectivity. In related work, complexes of sequence-specific DNA binding proteins, E. coli trp repressor, lac repressor and Eco RI methyl transferase with 2- thiouracil- or 2-thiothymine-containing oligomers will be studied. Triplet-triplet energy transfer from the sulfur-derivatized base of DNA to protein trp residues will be utilized to obtain structural information. Extensive use will be made of mutated proteins and enzymes that involve conservative substitution of intrinsic trp residues by other amino acids as an aid in interpretation of the spectroscopic results. Any information that we can provide that bears on the origins of sequence selectivity of p7 and p8, in particular, would have important implications in such areas as the design of novel HIV antiviral agents.

该研究项目的广泛长期目标是获得 有关蛋白质核酸相互作用的分子水平的信息。 特别重要的是我们对因素的理解的进一步 影响核酸序列选择性和稳定性 特定的配合物。蛋白质与核酸之间的相互作用是 细胞过程的基本组成部分，并且是必不可少的 用于哺乳动物等传染剂的组装和功能 逆转录病毒。拟议研究的主要目的是研究 1型人类核素蛋白的核酸复合物的 免疫缺陷病毒（导致艾滋病的传染剂），HIV-1 NC P7， 和猿猴免疫缺陷病毒，SIV NC P8。以及所有已知的 逆转录病毒NC蛋白，P7和P8包含锌指CCHC阵列 包括高度保守的芳族残基，例如TRP和PHE。这些Zn 手指阵列与特定的核酸结合有关 包装。我们的研究将集中于使用光学检测 P7和P8配合物的三胞胎状态磁共振（ODMR） 原子衍生的核酸以确定芳族是否堆叠 相互作用涉及结合。三胞胎状态零场 通过各种复合形成引起TRP诱导的分裂移位 核酸将用于定量评估 芳香的堆叠相互作用到复杂的稳定性，从而 对序列选择性的影响。这个目标将需要进一步 回甲霉素（DNA二元互化抗生素）类似物的研究 通过ODMR光谱作为堆叠诱导的序列选择性的模型。 在相关工作中，序列特异性DNA结合蛋白的复合物，E。 大肠杆菌TRP阻遏物，LAC阻遏物和Eco Ri甲基转移酶，具有2- 将研究硫核酸或2-含硫噻嗪的低聚物。 Triplet-Triplet能量从DNA的硫衍生物基础转移到 蛋白质TRP残基将用于获取结构信息。 大量使用将由涉及的突变蛋白质和酶制成 保守固有的TRP残基用其他氨基酸取代 有助于解释光谱结果。任何信息 我们可以根据序列选择性的起源来提供 尤其是P7和P8将在此类领域具有重要意义 作为新型HIV抗病毒药的设计。