Department of Chemistry, Stanford University, Stanford, California 94305-5080, United States.
J Am Chem Soc. 2011 Mar 23;133(11):4046-52. doi: 10.1021/ja110256c. Epub 2011 Feb 25.
Truncated green fluorescent protein (GFP) with the 11th β-strand removed is potentially interesting for bioconjugation, imaging, and the preparation of semisynthetic proteins with novel spectroscopic or functional properties. Surprisingly, the truncated GFP generated by removing the 11th strand, once refolded, does not reassemble with a synthetic peptide corresponding to strand 11 but does reassemble following light activation. The mechanism of this process has been studied in detail by absorption, fluorescence, and Raman spectroscopy. The chromophore in this refolded truncated GFP is found to be in the trans configuration. Upon exposure to light a photostationary state is formed between the trans and cis conformations of the chromophore, and only truncated GFP with the cis configuration of the chromophore binds the peptide. A kinetic model describing the light-activated reassembly of this split GFP is discussed. This unique light-driven reassembly is potentially useful for controlling protein-protein interactions.
截短型绿色荧光蛋白（GFP）去除第 11 条β-链后，对于生物偶联、成像以及制备具有新型光谱或功能特性的半合成蛋白可能很有意义。令人惊讶的是，通过去除第 11 条链生成的截短 GFP，一旦重新折叠，不会与对应于第 11 条链的合成肽重新组装，但在光激活后会重新组装。通过吸收、荧光和拉曼光谱详细研究了这个过程的机制。在这个重新折叠的截短 GFP 中，发色团被发现处于反式构型。暴露在光线下时，发色团的顺式和反式构象之间形成光稳定状态，只有具有顺式构象的发色团的截短 GFP 才能结合肽。讨论了描述该分裂 GFP 光激活重组装的动力学模型。这种独特的光驱动重组装对于控制蛋白质-蛋白质相互作用可能很有用。
