MOLECULAR RECOGNITION IN NUCLEAR PROTEIN TRANSPORT

核蛋白运输中的分子识别

• 批准号: 6519884
• 负责人: Robert F. Standaert
• 金额: $ 15.99万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-03-01 至 2005-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6519884
• 关键词: cell nucleus; intracellular transport; membrane permeability; membrane transport proteins; nuclear receptors; nucleoproteins; peptide chemical synthesis; photochemistry; protein localization; protein signal sequence; protein structure function; protein transport; receptor binding; technology /technique development

Nuclear protein import is a carefully orchestrated and regulated process; many proteins, notably transcription factors that activate genes selectively, are moved into the nucleus only in response to specific signals. Cell division, immune response, differentiation, development, viral infection and other important processes are intimately tied to the import process. Nuclear localization signals (NLSs), which direct nuclear entry of proteins, are short peptide motifs recognized by a specific, heterodimeric receptor (karyopherin ((/(). This proposal describes an approach to developing photo-regulated karyopherin ligands as tools for controlling protein localization in vivo with light. Novel, photochemically regulatable amino acids and carboxylic acids will be employed to generate mutants of native signals that bind karyopherin in a photo-regulated manner. The reagents are based on the azobenzene chromophore and fall into two groups: photoelastic amino acids that expand and contract the peptide backbone, and alpha, alpha-difluoro-phosphonomethyl azobenzenes which are intended to mimic reversible phosphorylation in response to light. The reagents will be applied in a potentially general strategy of systematic mutagenesis that targets native NLSs and proximal regulatory regions; principal targets include the SV40 T antigen NLS and the bipartite nucleoplasmin NLS. Mutagenesis will be implemented by efficient, parallel synthesis on the solid phase, and candidates will be screened on the solid phase for differential binding to karyopherin in response to light. Specific hypotheses to be tested are that the ability of two peptide domains to chelate the receptor can be controlled with photo-elastic amino acids, and that altered placement of a phosphate isostere by light can alter receptor-binding properties of the signals. Candidates displaying photosensitive receptor binding will be conjugated to protein carriers. The proteins will be introduced into permeabilized mammalian cells, and the effect of light on their intracellular localization will be determined. It is anticipated that signal-protein conjugates, introduced into living cells, would have light-dependent localization that could be used to control cellular processes such as gene transcription.

核蛋白输入是一个精心策划和监管的过程 过程;许多蛋白质，特别是激活基因的转录因子 仅响应特定信号才选择性地移入细胞核。 细胞分裂、免疫反应、分化、发育、病毒感染 以及其他重要流程与进口流程密切相关。 核定位信号（NLS），引导蛋白质进入核， 是被特定异二聚体受体识别的短肽基序 (karyopherin ((/())。该提案描述了一种开发方法 光调节核转运蛋白配体作为控制蛋白质的工具 用光进行体内定位。 新型光化学调节氨基酸和羧酸将 用于产生结合核传递蛋白的天然信号突变体 光调节方式。该试剂基于偶氮苯发色团 并分为两类：光弹性氨基酸，可膨胀和收缩 肽主链和 α, α-二氟膦酰甲基偶氮苯 旨在模拟响应光的可逆磷酸化。 这些试剂将应用于潜在的系统性通用策略 针对天然 NLS 和邻近调控区域的诱变；主要的 靶标包括 SV40 T 抗原 NLS 和二分核浆蛋白 NLS。 诱变将通过固体上的高效、平行合成来实施 阶段，候选者将在固相上进行筛选以进行差异化 响应光与核传递蛋白结合。待检验的具体假设 两个肽结构域螯合受体的能力可以是 由光弹性氨基酸控制，并且改变了 光的磷酸等排体可以改变受体的结合特性 信号。 显示光敏受体结合的候选物将缀合至 蛋白质载体。这些蛋白质将被引入透化哺乳动物体内 细胞，光对其细胞内定位的影响将是 决定。预计信号-蛋白缀合物被引入 活细胞将具有光依赖性定位，可用于 控制细胞过程，例如基因转录。

项目成果

Abc amino acids: design, synthesis, and properties of new photoelastic amino acids.

Abc 氨基酸：新型光弹性氨基酸的设计、合成和特性。

• DOI: 10.1021/jo060763q
• 发表时间: 2006
• 期刊: The Journal of organic chemistry
• 作者: Standaert,RobertF;Park,SeungBum
• 通讯作者: Park,SeungBum

Simple mimetics of a nuclear localization signal (NLS).

核定位信号（NLS）的简单模拟。

• DOI: 10.1021/ol034640t
• 发表时间: 2003
• 期刊: Organic letters.
• 作者: Park,SeungBum;Ho,ThaiH;Reedy,BrianM;Connolly,MichaelD;Standaert,RobertF
• 通讯作者: Standaert,RobertF

Asymmetric synthesis and translational competence of L-alpha-(1-cyclobutenyl)glycine.

• DOI: 10.1021/ol049026b
• 发表时间: 2004-09
• 期刊: Organic letters
• 影响因子: 5.2
• 作者: L. Jayathilaka;Mahua Deb;R. Standaert

A photoregulated ligand for the nuclear import receptor karyopherin alpha.

核输入受体核转运蛋白 α 的光调节配体。

• DOI: 10.1016/s0968-0896(01)00230-9
• 发表时间: 2001
• 期刊: Bioorganic & medicinal chemistry
• 影响因子: 3.5
• 作者: Park,SB;Standaert,RF
• 通讯作者: Standaert,RF