A hierarchic approach to the design of hexameric helical barrels
A hierarchic approach to the design of hexameric helical barrels
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DOI：
10.1016/s0022-2836(02)00233-4
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发表时间：
2002-05-24
影响因子：
5.6
通讯作者：
DeGrado, WF
中科院分区：
文献类型：
--
作者：
Ghirlanda, G;Lear, JD;DeGrado, WF
大分子组件的设计是一项对蛋白质工程和纳米技术都有影响的工作。采用层次化的方法设计了一种反平行的六面体螺旋管状组件。在以前的研究中，根据第一性原理设计了一个结构域互换的二聚体三螺旋纤维束。在晶格中，三个二聚体围绕一个3倍的旋转轴形成一个六面体组件。虽然在溶液中没有观察到这种六聚体组装，但可以通过将每个单体的三个极性残基改变为疏水残基(两个Phe和一个Trp)来稳定其形成。基于DSD(PDB加入码1G6U)的结晶学坐标的分子模型表明，这些侧链堆积在六角体束的中心空腔(超核)中。用分析超速离心法、荧光光谱学、圆二色谱和盐酸胍变性来确定六聚体的组装情况。为了探索稳定六聚体的要求，我们系统地改变了六聚体超核中一个Phe残基的极性和空间体积。根据这种侧链的性质，有可能以可预测的方式调节六角体的稳定性。这个六聚体蛋白质家族可能为构建蛋白质提供了一个有用的框架，这些蛋白质可以改变其低聚状态，以响应小分子的结合。(C)2002爱思唯尔科学有限公司。保留所有权利。
The design of large macromolecular assemblies is an endeavor with implications for protein engineering as well as nanotechnology. A hierarchic approach was used to design an antiparallel hexameric, tubular assembly of helices. In previous studies, a domain-swapped, dimeric three-helix bundle was designed from first principles. In the crystal lattice, three dimers associate around a 3-fold rotational axis to form a hexameric assembly. Although this hexameric assembly was not observed in solution, it was possible to stabilize its formation by changing three polar residues per monomer to hydrophobic (two Phe and one Trp) residues. Molecular models based on the crystallographic coordinates of DSD (PDB accession code 1G6U) show that these side-chains pack in the central cavity (the supercore) of the hexameric bundle. Analytical ultracentrifugation, fluorescence spectroscopy, CD spectroscopy, and guanidine-HCl denaturation were used to determine the assembly of the hexamer. To probe the requirements for stabilizing the hexamer, we systematically varied the polarity and steric bulk of one of the Phe residues in the supercore of the hexamer. Depending on the nature of this sidechain, it is possible to modulate the stability of the hexamer in a predictable manner. This family of hexameric proteins may provide a useful framework for the construction of proteins that change their oligomeric states in response to binding of small molecules. (C) 2002 Elsevier Science Ltd. All rights reserved.