Influence of Translation on Protein Folding

翻译对蛋白质折叠的影响

• 批准号: 7681116
• 负责人: Patricia Louise Clark
• 金额: $ 27.02万
• 依托单位: UNIVERSITY OF NOTRE DAME
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-09-01 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7681116
• 关键词: Affect; Algorithms; Amino Acid Sequence; Antibodies; Appearance; Bacteriophage P22; Binding; Cells; Codon Nucleotides; Coupled; Distant; Environment; Evolution; Fluorescence Anisotropy; Genetic Translation; Goals; High temperature of physical object; In Vitro; Kinetics; Laboratories; Length; Literature; Location; Mass Spectrum Analysis; Measurement; Measures; Messenger RNA; Methods; Modeling; Molecular Conformation; Muscle Rigidity; Nature; Peptide Sequence Determination; Peptides; Process; Proteins; Reporting; Research Personnel; Ribosomes; Site; Structure; Translations; aggregation pathway; base; design; in vivo; novel; novel strategies; polypeptide; preference; programs; protein folding; three dimensional structure

DESCRIPTION (provided by applicant): Successful protein folding involves the conversion of a linear polypeptide into a stable and biologically active 3D structure. This process has been studied for decades by denaturing purified, full-length polypeptides in vitro, diluting away the denaturant, and observing the refolding process. Careful measurements of refolding kinetics and populated partially folded conformations have revealed much about the refolding process. What is still lacking, however, is an understanding of how these in vitro results relate to protein folding in vivo. Protein folding in vivo is a fundamentally different process: In particular, the starting ensemble for folding in vivo is a growing nascent polypeptide chain, rather than a full-length chain. We have very little information about how this fundamentally different landscape for the growing chain relates to refolding mechanisms observed in vitro. Yet this difference may help explain why there are some native state topologies that are well represented in vivo, but difficult to refold in vitro. In this proposal, the influence of translation on protein folding in vivo will be studied from two perspectives: (1) The effects of translation pausing on folding yield. Rare codons can cause the ribosome to pause during translation, and literature reports have connected pauses with folding yield. For this proposal, novel methods have been developed to identify and modulate rare codon-derived translation pauses, and the effects on downstream folding. (2) The relationships between co-translational nascent chain conformations, in vitro refolding conformations, and the conformations of free, truncated polypeptides. Does protein refolding in vitro faithfully reproduce the conformations of nascent chains? For native structures, with contacts between distant portions of the polypeptide chain, this is unlikely. What alternative conformations are formed by these nascent chains? Are they also formed by free peptides? Results from in vitro refolding studies have revealed general principles for protein refolding; results from this proposal will be used to develop principles for protein folding in vivo.

描述（由申请人提供）：成功的蛋白质折叠涉及将线性多肽转化为稳定且具有生物活性的3D结构。数十年来，通过在体外贬低了纯化的，全长的多肽，稀释变性剂并观察重新折叠过程来研究这个过程数十年。仔细测量重折叠动力学和人口稠密的部分折叠构象已经揭示了有关重新折叠过程的很多东西。然而，仍然缺乏的是对这些体外结果如何与体内蛋白质折叠有关的理解。体内蛋白质折叠是一个根本不同的过程：尤其是在体内折叠的起始集合是一个生长的新生多肽链，而不是全长链。对于生长链的这种根本不同的景观，我们几乎没有信息与在体外观察到的重新折叠机制有关。然而，这种差异可能有助于解释为什么有一些本地状态拓扑在体内很好地表示，但很难在体外重新折叠。在此提案中，将从两个角度研究翻译对体内蛋白质折叠的影响：（1）平移暂停对折叠产量的影响。稀有密码子可以在翻译过程中导致核糖体停顿，文献报告已将停顿连接起来，折叠产量。对于此建议，已经开发出新的方法来识别和调节稀有密码子衍生的平移暂停，以及对下游折叠的影响。 （2）共转变的新生链构象，体外重折叠构象与游离，截短多肽的构象之间的关系。蛋白质在体外的重新折叠是否会忠实地再现新生链的构象？对于本地结构，在多肽链的遥远部分之间具有接触，这不太可能。这些新生的链形成了哪些替代构象？它们也由游离肽形成吗？体外重新折叠研究的结果揭示了蛋白质重折叠的一般原理。该提案的结果将用于制定体内蛋白质折叠的原理。