Iwane A H, Ikeda I, Kimura Y, Fujiyoshi Y, Altendorf K, Epstein W
Protein Engineering Research Institute, Suita, Osaka, Japan.
FEBS Lett. 1996 Nov 4;396(2-3):172-6. doi: 10.1016/0014-5793(96)01096-4.
A variant form of the Kdp-ATPase of Escherichia coli was overproduced to a level approaching 37% of the protein in the inner membrane of this organism. Membranes from overproducing cells were prepared with an inside-out orientation. Incubation of the membranes on ice for 1-2 weeks in the presence of sodium vanadate resulted in the formation of two-dimensional crystals of the Kdp-ATPase. The calculated projection map of the p1 crystal form showed three prominent density peaks at a resolution of 22 A. This technique is a useful and simple method to obtain low-resolution structures of membrane proteins.
大肠杆菌Kdp - ATP酶的一种变体形式被过量表达,其水平接近该生物体内膜中蛋白质的37%。过量表达细胞的膜以内外翻转的方向制备。在钒酸钠存在下,将膜在冰上孵育1 - 2周,导致形成Kdp - ATP酶的二维晶体。计算得到的p1晶型投影图在22埃分辨率下显示出三个明显的密度峰。该技术是获得膜蛋白低分辨率结构的一种有用且简单的方法。