Tommassen J, van der Ley P, van Zeijl M, Agterberg M
EMBO J. 1985 Jun;4(6):1583-7. doi: 10.1002/j.1460-2075.1985.tb03820.x.
The genes ompC and phoE of Escherichia coli K-12 encode outer membrane pore proteins that are very homologous. To study the structure-function relationship of these proteins, we have constructed a series of ompC-phoE hybrid genes in which the DNA encoding part of one protein is replaced by the corresponding part of the other gene. These hybrid genes were easily obtained by using in vivo recombination. The fusion sites in the hybrid genes were localized by restriction enzyme mapping. The hybrid gene products were normally expressed and they were characterized with respect to functions and properties in which the native OmpC and PhoE proteins differ, such as pore characteristics, the receptor activity for phages and the binding of specific antibodies. Three regions within the N-terminal 130 amino acids were localized which determine pore characteristics and a segment between residues 75 and 110 contains amino acids which determine specificity for PhoE phages. A major cell surface-exposed region is located between residues 142 and 267. This region contains residues which are required for the binding of monoclonal antibodies directed against the cell surface-exposed part of PhoE and residues which determine specificity for OmpC phages.
大肠杆菌K-12的ompC和phoE基因编码非常同源的外膜孔蛋白。为了研究这些蛋白质的结构-功能关系，我们构建了一系列ompC-phoE杂交基因，其中一个蛋白质编码部分的DNA被另一个基因的相应部分所取代。这些杂交基因通过体内重组很容易获得。通过限制性酶切图谱确定了杂交基因中的融合位点。杂交基因产物正常表达，并就天然OmpC和PhoE蛋白不同的功能和特性进行了表征，如孔特征、噬菌体受体活性和特异性抗体的结合。确定了N端130个氨基酸内的三个区域决定孔特征，75至110位残基之间的一段含有决定对PhoE噬菌体特异性的氨基酸。一个主要的细胞表面暴露区域位于142至267位残基之间。该区域包含针对PhoE细胞表面暴露部分的单克隆抗体结合所需的残基以及决定对OmpC噬菌体特异性的残基。
