OPTIMALLY STABLE PROTEINS FROM PEPTIDE LIBRARIES

来自肽库的最佳稳定蛋白质

• 批准号: 7725265
• 负责人: MARTIN A CASE
• 金额: $ 3.93万
• 依托单位: UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-01 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7725265
• 关键词: Amino Acid Sequence; Chemicals; Chemistry; Computer Retrieval of Information on Scientific Projects Database; DNA; Disruption; Elements; Funding; Grant; In Vitro; Institution; Kinetics; Ligands; Metals; Modification; Peptide Library; Peptides; Pharmaceutical Preparations; Protein Overexpression; Proteins; Proteolysis; Research; Research Personnel; Resistance; Resources; Source; Structure; Surface; Therapeutic; Thermodynamics; United States National Institutes of Health; base; design; globular protein; immunogenicity; in vivo; interest; protein protein interaction; small molecule

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The long-term objective of this application is to design proteins that can be selected ex-vivo for their high stability. Such proteins will be resistant to proteolysis, and will tolerate surface modifications that will reduce their immunogenicity. As such, they will be ideal candidates for therapeutic applications requiring the disruption of endogenous protein-protein interactions. Successful completion of the proposed research will reveal amino-acid sequences that confer optimal folding stability on small globular proteins. An in vitro chemical assembly and selection strategy provides candidate sequences for subsequent in vivo expression. Specifically, assembly of peptide secondary structural elements into a pre-determined fold is accomplished using exchange-labile metal-ligand chemistry. The subunits exchange partners under thermodynamic control, and the most stable associations are isolated and their amino acid sequences determined. DNA cassettes are constructed based on the sequence information returned from the in vitro selection using synthetic trinucleotides. Overexpressed proteins are isolated and characterized. Of particular interest are the folding stabilities of these de novo proteins, and their folding kinetics as compared with natural proteins of similar structure. The creation of proteins with extremely stable folded structures will open the door to protein-based drugs for applications in therapeutic strategies where small molecules are ineffective.

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 这一应用的长期目标是设计能够在体外选择的蛋白质，因为它们具有高度的稳定性。这样的蛋白质将抵抗蛋白质分解，并将容忍降低其免疫原性的表面修饰。因此，它们将是需要破坏内源性蛋白质-蛋白质相互作用的治疗应用的理想候选者。这项拟议研究的成功完成将揭示赋予小球状蛋白最佳折叠稳定性的氨基酸序列。体外化学组装和选择策略为随后的体内表达提供了候选序列。具体地说，利用交换不稳定的金属配体化学来完成将多肽二级结构元件组装到预定折叠中。亚基在热力学控制下交换伙伴，最稳定的结合被分离出来，并确定它们的氨基酸序列。 DNA盒是基于使用合成的三核苷酸从体外选择返回的序列信息构建的。对过表达的蛋白质进行分离和鉴定。尤其令人感兴趣的是这些从头蛋白质的折叠稳定性，以及它们与结构相似的天然蛋白质相比的折叠动力学。具有极其稳定的折叠结构的蛋白质的产生将为以蛋白质为基础的药物在小分子无效的治疗策略中的应用打开大门。