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Backbone Effects on Protein Stability and Folding

主链对蛋白质稳定性和折叠的影响

基本信息

批准号：
8066833
负责人：
PHILIP E DAWSON
金额：
$ 9.83万
依托单位：
SCRIPPS RESEARCH INSTITUTE, THE
依托单位国家：
美国
项目类别：
财政年份：
1999
资助国家：
美国
起止时间：
1999-05-01 至 2011-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8066833
关键词：
Acylphosphatase Address Amino Acids Aminoisobutyric Acids Biochemical Biological Models Complement Complex Computer Analysis Crystallography Data Development Elements Esters Fluorescence Functional RNA Generations Glycine Human Hydrogen Bonding Hydroxy Acids Individual Kinetics Leucine Zippers Ligation Measurement Measures Methods Modeling Modification Molecular Molecular Conformation Molecular Structure Muscle Mutagenesis Mutation Nature Peptides Phase Phosphoric Monoester Hydrolases Process Property Protein Biosynthesis Protein Engineering Proteins Relative (related person)Research Research Personnel Role Science Series Side Site Site-Directed Mutagenesis Solid Solvents Source System Techniques Tertiary Protein Structure Testing Thermodynamics Variant Vertebral column Work alpha helix analog analytical ultracentrifugation base beta pleated sheet chemical synthesis chymotrypsin inhibitor combinatorial chemistry computer studies insight interest mutant novel peptide chemical synthesis polypeptide programs protein folding protein structure synthetic protein three dimensional structure tool trend

项目摘要

The broad objective of this research program is to understand the relationship between the molecular structure of a polypeptide chain and its ability to fold into a defined, three-dimensional structure. Most studies on protein folding and stability have focused on the role of amino acid side-chains using site-directed mutagenesis. We propose to diverge from this trend by using the total synthesis of proteins to chemically modify the polypeptide backbone. We believe that systematic variation of the backbone will give insight into the fundamental forces that stabilize proteins and the processes through which they fold. We have demonstrated thatjpha-hydroxy acids can be incorporated into proteins in a site-specific manner using peptide synthesis and chemical ligation methods. We have utilized this modification to analyze the energetic contributions of specific hydrogen bonds in the GCN4 coiled coil and, in kinetic studies, the folding transition state of the chymotrypsin inhibitor CI2. These studies indicate that backbone modification provides direct information on the formation of backbone hydrogen bonding in the native state and folding transition state ensemble that is not observed using traditional side chain mutagenesis methods. This proposal aims to answer specific questions regarding the folding transition states of GCN4 and CI2 and to extend these studies to the B1 domain of Protein L and acylphosphatase. These proteins have been selected to take advantage of previous work using site directed mutagenesis to analyze the folding transitions of these proteins. In addition, recent computational analyses of these proteins have made specific predictions about folding that cannot be addressed by traditional experimental mutagenesis strategies. We feel that this use of non-coded modifications such as ester bonds for thermodynamic and kinetic measurements of protein folding will enable new insights into the molecular basis of protein folding and stability and provide data for the continued development of computational approaches to this problem.

这项研究计划的主要目标是了解分子之间的关系，多肽链的结构及其折叠成确定的三维结构的能力。大多数研究关于蛋白质折叠和稳定性的研究主要集中在氨基酸侧链的作用上，诱变我们建议偏离这一趋势，通过使用蛋白质的全合成，修饰多肽骨架。我们相信，脊柱的系统性变异将使我们了解稳定蛋白质的基本力量以及它们折叠的过程。我们有证明了α-羟基酸可以以位点特异性的方式掺入蛋白质中，肽合成和化学连接方法。我们已经利用这种修改来分析能量 GCN 4卷曲螺旋中特定氢键的贡献，以及动力学研究中的折叠转变胰凝乳蛋白酶抑制剂CI 2的状态。这些研究表明，骨架修饰提供了直接的关于在天然状态和折叠过渡状态下骨架氢键形成的信息这是使用传统侧链诱变方法未观察到的系综。该提案旨在回答关于GCN 4和Cl 2的折叠过渡态的具体问题，将这些研究扩展到蛋白L和酰基磷酸酶的B1结构域。这些蛋白质选择利用以前的工作，使用定点诱变来分析折叠这些蛋白质的转变。此外，最近对这些蛋白质的计算分析已经使特定的传统实验突变策略无法解决的折叠预测。我们我觉得这种使用非编码修饰如酯键的热力学和动力学蛋白质折叠的测量将使人们对蛋白质折叠的分子基础有新的认识，稳定性，并提供数据的计算方法，这个问题的持续发展。

项目成果

期刊论文数量（29）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Interactions between redox complexes and semiconductor quantum dots coupled via a peptide bridge.

DOI：
10.1021/ja805456x
发表时间：
2008-12-10
期刊：
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
影响因子：
15
作者：
Medintz, Igor L.;Pons, Thomas;Trammell, Scott A.;Grimes, Amy F.;English, Doug S.;Blanco-Canosa, Juan B.;Dawson, Philip E.;Mattoussi, Hedi
通讯作者：
Mattoussi, Hedi

N-methylcysteine-mediated total chemical synthesis of ubiquitin thioester.

DOI：
10.1039/c000332h
发表时间：
2010-05-21
期刊：
Organic & biomolecular chemistry
影响因子：
3.2
作者：
Erlich LA;Kumar KS;Haj-Yahya M;Dawson PE;Brik A
通讯作者：
Brik A

Traceless ligation of cysteine peptides using selective deselenization.

DOI：
10.1002/anie.201001900
发表时间：
2010-09-17
期刊：
ANGEWANDTE CHEMIE-INTERNATIONAL EDITION
影响因子：
16.6
作者：
Metanis, Norman;Keinan, Ehud;Dawson, Philip E.
通讯作者：
Dawson, Philip E.

High-efficiency labeling of sialylated glycoproteins on living cells.

DOI：
10.1038/nmeth.1305
发表时间：
2009-03
期刊：
NATURE METHODS
影响因子：
48
作者：
Zeng, Ying;Ramya, T. N. C.;Dirksen, Anouk;Dawson, Philip E.;Paulson, James C.
通讯作者：
Paulson, James C.

Bisaryl hydrazones as exchangeable biocompatible linkers.