Zukin R S, Klos M F, Hirsch R E
Biophys J. 1986 Jun;49(6):1229-35. doi: 10.1016/S0006-3495(86)83752-3.
The Salmonella typhimurium periplasmic histidine-binding J-protein is one of four proteins encoded by the histidine transport operon. Mutant J-protein hisJ5625 binds L-histidine, but does not transport it. The tertiary structure and conformational dynamics of native and mutant J-protein have been compared using steady state fluorescence, fluorescence polarization, and fluorescence energy transfer measurements. The two proteins have different three-dimensional structures and exhibit different responses to histidine binding. Ligand-induced conformational changes were demonstrated in both J-proteins using fluorescence energy transfer (distant reporter method) between the single tryptophan residue per mole of protein and a fluorescein-labeled methionine residue. However, the conformational change of the mutant protein is qualitatively and quantitatively different from that of the wild-type protein. Moreover, the microenvironment of the tryptophan and its distance from the labeled methionine (44A for the wild type, 60A for the mutant J-protein) are different in the two proteins. In conclusion, these results indicate that the specific conformational change induced in the wild type J-protein is a necessary requirement for the transport of L-histidine.
鼠伤寒沙门氏菌周质组氨酸结合J蛋白是组氨酸转运操纵子编码的四种蛋白之一。突变型J蛋白hisJ5625能结合L-组氨酸，但不能转运它。已使用稳态荧光、荧光偏振和荧光能量转移测量方法比较了天然和突变型J蛋白的三级结构和构象动力学。这两种蛋白具有不同的三维结构，并且对组氨酸结合表现出不同的响应。使用每摩尔蛋白中的单个色氨酸残基与荧光素标记的甲硫氨酸残基之间的荧光能量转移（远距离报告法），在两种J蛋白中均证实了配体诱导的构象变化。然而，突变蛋白的构象变化在定性和定量上均与野生型蛋白不同。此外，两种蛋白中色氨酸的微环境及其与标记甲硫氨酸的距离（野生型为44埃，突变型J蛋白为60埃）也不同。总之，这些结果表明，野生型J蛋白中诱导的特定构象变化是L-组氨酸转运的必要条件。
