Corn Jacob E, Berger James M
Department of Molecular and Cell Biology, University of California, Berkeley, 327B Hildebrand Hall, Berkeley, CA 94720, USA.
Structure. 2007 Jul;15(7):773-80. doi: 10.1016/j.str.2007.05.006.
Structural studies of macromolecular complexes have produced extraordinary insights into a wide variety of biological processes. Unfortunately, as structural biologists pursue larger and more challenging assemblies, weakly stable and/or nonspecific interactions can become significant roadblocks to structure determination. We have developed a rapid and effective pool-based screen, termed FASTDXL (focused array screening technique for disulfide X-linking), to produce and identify disulfide-stabilized protein-nucleic acid assemblies. A significant strength of FASTDXL is that it can take advantage of prior structural knowledge about molecular interactions, but does not necessarily rely upon it. A detailed application of the approach to the difficult problem of trapping a bacterial primase-ssDNA complex is described, validating the method as a route toward obtaining diffracting crystals suitable for structure determination.
大分子复合物的结构研究为各种各样的生物过程带来了非凡的见解。不幸的是，随着结构生物学家研究更大、更具挑战性的组装体，弱稳定性和/或非特异性相互作用可能成为结构测定的重大障碍。我们开发了一种基于池的快速有效筛选方法，称为FASTDXL（用于二硫键交联的聚焦阵列筛选技术），以产生并鉴定二硫键稳定的蛋白质-核酸组装体。FASTDXL的一个显著优势在于它可以利用关于分子相互作用的先前结构知识，但不一定依赖于此。本文描述了该方法在捕获细菌引发酶-单链DNA复合物这一难题上的详细应用，验证了该方法是获得适合结构测定的衍射晶体的一条途径。
