北京大学理论生物学中心/化学与分子工程学院来鲁华教授领导的课题组在功能蛋白质设计方面取得可喜进展，其研究成果发表于3月27日出版的《美国科学院院刊》上（Liu S, Liu SY, Zhu XL, Liang HH, Cao AN, Chang ZJ and Lai LH*. Nonnatural protein-protein interaction-pair design by key residues grafting. PNAS,104:5330-5335,2007）。论文发表后引起了学界关注，BioCentury就此成果发表了专访。该论文的第一作者刘森同学是理论生物学中心2002年正式招收的第一批跨学科的研究生。
蛋白质在生命活动中占有非常重要的地位，几乎所有的生命活动都需要蛋白质的参与。生命现象纷繁复杂，所涉及到的蛋白质功能各异。在履行生命功能的过程中，蛋白质不断地与其它分子（如蛋白质，核酸，小分子等）发生相互作用。就如同现实生活中，我们看到的钥匙有各种各样的形状，但是只要它们的关键部分“齿纹”一样，就能够有同样的功能，打开同一把锁。相互作用的两个蛋白质，就有如“锁”和“钥匙”的关系。来鲁华教授课题组的研究目的，就是将一个蛋白质行使功能的关键“齿纹”（氨基酸残基），复制（“嫁接”）到另一个结构完全不同的蛋白质上，使它具有开同一把“锁”的功能（和同一个蛋白质结合）。
蛋白质和蛋白质结合时，往往存在少数几个非常关键的残基，对结合起到主要作用。基于这种情况，来鲁华教授课题组发展了一种“蛋白质关键残基嫁接”的算法，用于将一个蛋白质的关键功能性残基嫁接到另一个不同结构的蛋白质上，并将这种方法应用到了实际体系的研究中，取得了成功。促红细胞生成素（EPO）通过和它的受体（EPOR）相互作用，促进红细胞的分化和成熟。将EPO上的关键功能性残基嫁接到一个结构完全不同的PH结构域蛋白上后，PH蛋白具有了和EPOR结合的功能，而这种功能在自然界中是不存在的。这种方法将有可能应用在更为广泛的例子上，实现蛋白质功能的自由设计。
原始出处:
Published online before print March 19, 2007, 10.1073/pnas.0606198104
PNAS | March 27, 2007 | vol. 104 | no. 13 | 5330-5335
Nonnatural protein–protein interaction-pair design by key residues grafting
Sen Liu*,, Shiyong Liu*,, Xiaolei Zhu*, Huanhuan Liang*, Aoneng Cao*, Zhijie Chang, and Luhua Lai*,,
*Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China; Center for Theoretical Biology, Peking University, Beijing 100871, China; and Department of Biological Sciences and Biotechnology, School of Medicine, Tsinghua University, Beijing 100084, China
Edited by Stephen L. Mayo, California Institute of Technology, Pasadena, CA, and approved January 22, 2007 (received for review July 21, 2006)
Protein–protein interface design is one of the most exciting fields in protein science; however, designing nonnatural protein–protein interaction pairs remains difficult. In this article we report a de novo design of a nonnatural protein–protein interaction pair by scanning the Protein Data Bank for suitable scaffold proteins that can be used for grafting key interaction residues and can form stable complexes with the target protein after additional mutations. Using our design algorithm, an unrelated protein, rat PLC1-PH (pleckstrin homology domain of phospholipase C-1), was successfully designed to bind the human erythropoietin receptor (EPOR) after grafting the key interaction residues of human erythropoietin binding to EPOR. The designed mutants of rat PLC1-PH were expressed and purified to test their binding affinities with EPOR. A designed triple mutation of PLC1-PH (ERPH1) was found to bind EPOR with high affinity (KD of 24 nM and an IC50 of 5.7 µM) both in vitro and in a cell-based assay, respectively, although the WT PLC1-PH did not show any detectable binding under the assay conditions. The in vitro binding affinities of the PLC1-PH mutants correlate qualitatively to the computational binding affinities, validating the design and the protein–protein interaction model. The successful practice of finding a proper protein scaffold and making it bind with EPOR demonstrates a prospective application in protein engineering targeting protein–protein interfaces.
de novo design of protein–protein interaction pair | erythropoietin | functional site grafting | key residue at interface
