Banatao D Rey, Cascio Duilio, Crowley Christopher S, Fleissner Mark R, Tienson Heather L, Yeates Todd O
California Nanosystems Institute, University of California, Boyer Hall 259, P.O. Box 951570, Los Angeles, CA 90095-1570, USA.
Proc Natl Acad Sci U S A. 2006 Oct 31;103(44):16230-5. doi: 10.1073/pnas.0607674103. Epub 2006 Oct 18.
Previous studies of symmetry preferences in protein crystals suggest that symmetric proteins, such as homodimers, might crystallize more readily on average than asymmetric, monomeric proteins. Proteins that are naturally monomeric can be made homodimeric artificially by forming disulfide bonds between individual cysteine residues introduced by mutagenesis. Furthermore, by creating a variety of single-cysteine mutants, a series of distinct synthetic dimers can be generated for a given protein of interest, with each expected to gain advantage from its added symmetry and to exhibit a crystallization behavior distinct from the other constructs. This strategy was tested on phage T4 lysozyme, a protein whose crystallization as a monomer has been studied exhaustively. Experiments on three single-cysteine mutants, each prepared in dimeric form, yielded numerous novel crystal forms that cannot be realized by monomeric lysozyme. Six new crystal forms have been characterized. The results suggest that synthetic symmetrization may be a useful approach for enlarging the search space for crystallizing proteins.
先前关于蛋白质晶体对称性偏好的研究表明，对称蛋白质，如同源二聚体，平均而言可能比不对称的单体蛋白质更容易结晶。天然单体的蛋白质可以通过在诱变引入的单个半胱氨酸残基之间形成二硫键而人工制成同源二聚体。此外，通过创建各种单半胱氨酸突变体，可以为给定的目标蛋白质生成一系列不同的合成二聚体，预计每个二聚体都能从其增加的对称性中获益，并表现出与其他构建体不同的结晶行为。该策略在噬菌体T4溶菌酶上进行了测试，这是一种作为单体结晶已被详尽研究的蛋白质。对三个均以二聚体形式制备的单半胱氨酸突变体进行的实验产生了许多单体溶菌酶无法实现的新型晶体形式。已对六种新晶体形式进行了表征。结果表明，合成对称化可能是扩大蛋白质结晶搜索空间的一种有用方法。
