SPECTROSCOPY OF HEAVY ATOM-PERTURBED BIOPOLYMERS

重原子扰动生物聚合物的光谱学

• 批准号: 2518618
• 负责人: AUGUST H MAKI
• 金额: $ 13.72万
• 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 1998-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2518618
• 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.

这项研究项目的长期目标是获得 关于蛋白质-核酸相互作用的分子水平的信息。 尤其重要的是加深我们对各种因素的理解。 影响核酸序列的选择性和稳定性 特定的复合体。蛋白质和核酸之间的相互作用是 是细胞过程的基本组成部分，也是必不可少的 用于感染病原体的组装和功能，如哺乳动物 逆转录病毒。拟议研究的主要具体目的是研究 人类1型核衣壳蛋白核酸复合体的研究 免疫缺陷病毒(一种导致艾滋病的感染剂)，HIV-1NC p7， 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， 包括高度保守的芳香族残基，如色氨酸和苯丙氨酸。这些锌 手指阵列与特定的核酸结合和 包装。我们的研究将集中在光学检测的使用上 P7和p8重离子配合物的三重态磁共振(ODMR) 原子衍生核酸确定芳香族堆积是否 相互作用涉及到绑定。三重态零场 不同结构的络合物在色氨酸中引起的分裂位移 核酸将被用来定量评估 芳香族堆积相互作用对络合物稳定性的影响，从而使其 对序列选择性的影响。这一目标将需要进一步 棘霉素(DNA双嵌入型抗生素)类似物的研究 通过ODMR光谱作为堆积诱导序列选择性的模型。 在相关工作中，序列特异性DNA结合蛋白、E. ColiTrp阻遏物、Lac阻遏物和Eco RI甲基转移酶与2- 将研究含有硫尿嘧啶或2-硫代胸腺嘧啶的低聚物。 DNA硫衍生物碱基的三重态-三重态能量转移 蛋白质色氨酸残基将被用来获得结构信息。 将广泛使用突变的蛋白质和酶，涉及 其他氨基酸对固有色氨酸残基的保守替代 作为解释光谱结果的辅助工具。任何信息 我们可以提供与序列选择性的起源有关的信息 特别是p7和p8将在这些领域产生重要影响。 作为新型HIV抗病毒药物的设计。