通过铰链环诱变了解c-Src SH3结构域交换。

IF 3.8 4区 生物学 Q2 BIOCHEMICAL RESEARCH METHODS
M Carmen Salinas-Garcia, Marina Plaza-Garrido, Jose C Martinez, Ana Camara-Artigas
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引用次数: 0

摘要

从生物物理和结构的角度来看,c-Src SH3结构域是最具特征的模块化结构域之一。这个SH3结构域显示非规范的替代折叠,形成3D结构域交换的低聚物和淀粉样原纤维。这些特征使这种小蛋白质成为研究这些现象的理想模型。在进行三维结构域交换的蛋白质中,对有利于单体展开的区域和铰链环中的残基进行了深入研究。为了研究这些残基在c-Src SH3结构域展开中的作用,我们通过在c-Src SH3和Abl SH3结构域之间交换RT和n-Src环中的残基构建了几种嵌合蛋白。RT (β1和β2之间的区域)和n-Src (β2和β3之间的区域)环形成浅疏水槽的两侧,富含脯氨酸的基序序列与SH3结构域结合。除了结构信息外,我们还对这些嵌合结构进行了生物物理表征。承载Abl SH3环的c-Src SH3结构域在稳定性上表现出微小的变化。有趣的是,这些替换并不能阻止结构域交换二聚体的形成。然而,Abl SH3结构域内一个或两个环的交换会显著降低其稳定性,但不会促进3D结构域交换低聚物的形成。因此,我们的研究结果表明,虽然铰链环的组成可能在结构元素的交换中发挥作用,形成相互缠绕的二聚体,但它不是形成二聚体的唯一驱动力。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Understanding domain swapping in the c-Src SH3 domain through hinge-loop mutagenesis.

The c-Src SH3 domain is one of the best-characterized modular domains from a biophysical and structural point of view. This SH3 domain displays noncanonical alternative folding, forming 3D domain-swapped oligomers and amyloid fibrils. These features make this small protein an ideal model for studying these phenomena. Residues in the regions that favour unfolding of the monomer and those in the hinge loop have been deeply studied in proteins undergoing 3D domain swapping. To study the role of these residues in the unfolding of the c-Src SH3 domain, we have constructed several chimeric proteins by interchanging residues in the RT and n-Src loops between the c-Src SH3 and Abl SH3 domains. The RT (the region between β1 and β2) and n-Src (the region between β2 and β3) loops create two sides of the shallow hydrophobic groove where proline-rich motif sequences bind to the SH3 domain. In addition to the structural information, we have performed a biophysical characterization of these chimeric constructs. The c-Src SH3 domain bearing the loops of the Abl SH3 shows minor changes in stability. Interestingly, these replacements do not prevent the formation of domain-swapped dimers. However, the interchange of one or two loops within the Abl SH3 domain produces a noticeable reduction in its stability but does not promote the formation of 3D domain-swapped oligomers. Thus, our results indicate that although the composition of the hinge loop is likely to play a role in the interchange of structural elements to form the intertwined dimers, it is not the sole driving force in their formation.

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来源期刊
Acta Crystallographica. Section D, Structural Biology
Acta Crystallographica. Section D, Structural Biology BIOCHEMICAL RESEARCH METHODSBIOCHEMISTRY &-BIOCHEMISTRY & MOLECULAR BIOLOGY
CiteScore
4.50
自引率
13.60%
发文量
216
期刊介绍: Acta Crystallographica Section D welcomes the submission of articles covering any aspect of structural biology, with a particular emphasis on the structures of biological macromolecules or the methods used to determine them. Reports on new structures of biological importance may address the smallest macromolecules to the largest complex molecular machines. These structures may have been determined using any structural biology technique including crystallography, NMR, cryoEM and/or other techniques. The key criterion is that such articles must present significant new insights into biological, chemical or medical sciences. The inclusion of complementary data that support the conclusions drawn from the structural studies (such as binding studies, mass spectrometry, enzyme assays, or analysis of mutants or other modified forms of biological macromolecule) is encouraged. Methods articles may include new approaches to any aspect of biological structure determination or structure analysis but will only be accepted where they focus on new methods that are demonstrated to be of general applicability and importance to structural biology. Articles describing particularly difficult problems in structural biology are also welcomed, if the analysis would provide useful insights to others facing similar problems.
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