Dahiyat B I, Sarisky C A, Mayo S L
Division of Chemistry and Chemical Engineering, California Institue of Technology, Pasadena, 91125, USA.
J Mol Biol. 1997 Nov 7;273(4):789-96. doi: 10.1006/jmbi.1997.1341.
Several groups have applied and experimentally tested systematic, quantitative methods to protein design with the goal of developing general design algorithms. We have sought to expand the range of computational protein design by developing quantitative design methods for residues of all parts of a protein: the buried core, the solvent exposed surface, and the boundary between core and surface. Our goal is an objective, quantitative design algorithm that is based on the physical properties that determine protein structure and stability and which is not limited to specific folds or motifs. We chose the betabetaalpha motif typified by the zinc finger DNA binding module to test our design methodology. Using previously published sequence scoring functions developed with a combined experimental and computational approach and the Dead-End Elimination theorem to search for the optimal sequence, we designed 20 out of 28 positions in the test motif. The resulting sequence has less than 40% homology to any known sequence and does not contain any metal binding sites or cysteine residues. The resulting peptide, pda8d, is highly soluble and monomeric and circular dichroism measurements showed it to be folded with a weakly cooperative thermal unfolding transition. The NMR solution structure of pda8d was solved and shows that it is well-defined with a backbone ensemble rms deviation of 0. 55 A. Pda8d folds into the desired betabetaalpha motif with well-defined elements of secondary structure and tertiary organization. Superposition of the pda8d backbone to the design target is excellent, with an atomic rms deviation of 1.04 A.
几个研究小组已经应用并通过实验测试了系统的定量方法来进行蛋白质设计，目的是开发通用的设计算法。我们试图通过为蛋白质所有部分的残基开发定量设计方法来扩大计算蛋白质设计的范围：埋藏核心、溶剂暴露表面以及核心与表面之间的边界。我们的目标是一种客观的、基于决定蛋白质结构和稳定性的物理性质的定量设计算法，并且不限于特定的折叠或基序。我们选择了以锌指DNA结合模块为代表的ββα基序来测试我们的设计方法。使用先前通过实验和计算相结合的方法开发的序列评分函数以及死端消除定理来搜索最佳序列，我们在测试基序的28个位置中设计了20个。所得序列与任何已知序列的同源性小于40%，并且不包含任何金属结合位点或半胱氨酸残基。所得肽pda8d高度可溶且为单体，圆二色性测量表明它以弱协同热解折叠转变折叠。解决了pda8d的NMR溶液结构，结果表明它具有良好的定义，主链系综均方根偏差为0.55埃。Pda8d折叠成所需的ββα基序，具有明确的二级结构和三级结构元素。pda8d主链与设计目标的叠加效果极佳，原子均方根偏差为1.04埃。
