Perozzo M A, Ward K B, Thompson R B, Ward W W
Laboratory for the Structure of Matter, Naval Research Laboratory, Washington, D.C. 20375-5000.
J Biol Chem. 1988 Jun 5;263(16):7713-6.
The energy transfer protein, green fluorescent protein, from the hydromedusan jellyfish Aequorea victoria has been crystallized in two morphologies suitable for x-ray diffraction analysis. Hexagonal plates have been obtained in the P6122 or P6522 space group with a = b = 77.5, c = 370 A, and no more than three molecules per asymmetric unit. Monoclinic parallel-epipeds have been obtained in the C2 space group with a = 93.3, b = 66.5, c = 45.5 A, beta = 108 degrees, and one molecule per asymmetric unit. The monoclinic form is better suited for use in a structure determination, and a data set was collected from the native crystal. Time-resolved fluorescence measurements of large single crystals are possible due to the unique, covalently bound chromophore present in this molecule. Fluorescence emission spectra of Aequorea green fluorescent protein in solution and from either the hexagonal or monoclinic single crystal show similar profiles suggesting that the conformations of protein in solution and in the crystal are similar. Multifrequency phase fluorimetric data obtained from a single crystal were best fit by a single fluorescence lifetime very close to that exhibited by the protein in solution. The complementary structural data obtained from fluorescence spectroscopy and x-ray diffraction crystallography will aid in the elucidation of this novel protein's structure-function relationship.
来自水螅水母维多利亚多管水母的能量转移蛋白——绿色荧光蛋白，已结晶成两种适合X射线衍射分析的形态。在P6122或P6522空间群中获得了六边形板状晶体，a = b = 77.5，c = 370 Å，每个不对称单元不超过三个分子。在C2空间群中获得了单斜平行六面体晶体，a = 93.3，b = 66.5，c = 45.5 Å，β = 108°，每个不对称单元有一个分子。单斜晶型更适合用于结构测定，并且从天然晶体收集了数据集。由于该分子中存在独特的共价结合发色团，因此可以对大的单晶进行时间分辨荧光测量。溶液中的维多利亚多管水母绿色荧光蛋白以及六边形或单斜单晶的荧光发射光谱显示出相似的轮廓，这表明溶液中和晶体中蛋白质的构象相似。从单晶获得的多频相荧光数据通过非常接近溶液中蛋白质所表现出的单荧光寿命得到了最佳拟合。从荧光光谱学和X射线衍射晶体学获得的互补结构数据将有助于阐明这种新型蛋白质的结构-功能关系。
