Phizicky E M, Fields S
Department of Biochemistry, University of Rochester Medical School, New York 14642.
Microbiol Rev. 1995 Mar;59(1):94-123. doi: 10.1128/mr.59.1.94-123.1995.
The function and activity of a protein are often modulated by other proteins with which it interacts. This review is intended as a practical guide to the analysis of such protein-protein interactions. We discuss biochemical methods such as protein affinity chromatography, affinity blotting, coimmunoprecipitation, and cross-linking; molecular biological methods such as protein probing, the two-hybrid system, and phage display: and genetic methods such as the isolation of extragenic suppressors, synthetic mutants, and unlinked noncomplementing mutants. We next describe how binding affinities can be evaluated by techniques including protein affinity chromatography, sedimentation, gel filtration, fluorescence methods, solid-phase sampling of equilibrium solutions, and surface plasmon resonance. Finally, three examples of well-characterized domains involved in multiple protein-protein interactions are examined. The emphasis of the discussion is on variations in the approaches, concerns in evaluating the results, and advantages and disadvantages of the techniques.
蛋白质的功能和活性常常受到与其相互作用的其他蛋白质的调节。本综述旨在作为分析此类蛋白质-蛋白质相互作用的实用指南。我们讨论了生化方法，如蛋白质亲和色谱法、亲和印迹法、免疫共沉淀法和交联法；分子生物学方法，如蛋白质探测法、双杂交系统和噬菌体展示法；以及遗传学方法，如分离基因外抑制子、合成突变体和非连锁非互补突变体。接下来，我们描述了如何通过蛋白质亲和色谱法、沉降法、凝胶过滤法、荧光法、平衡溶液的固相采样法和表面等离子体共振法等技术来评估结合亲和力。最后，研究了涉及多种蛋白质-蛋白质相互作用的三个特征明确的结构域实例。讨论的重点在于方法的差异、评估结果时的注意事项以及这些技术的优缺点。
