Fuchs R, Jauris S, Lottspeich F, Preac-Mursic V, Wilske B, Soutschek E
Max von Pettenkofer-Institut, University of Munich, Germany.
Mol Microbiol. 1992 Feb;6(4):503-9. doi: 10.1111/j.1365-2958.1992.tb01495.x.
We describe the cloning and expression of the pc gene which encodes a major immunodominant protein of Borrelia burgdorferi, the causative agent of Lyme borreliosis. The pC protein was purified from lysates of B. burgdorferi strain PKo. After tryptic digestion of the pC protein the resulting oligopeptides were applied to a gas-phase sequenator. Thus partial amino acid sequences were obtained. The deduced oligonucleotides were used as hybridization probes. After Southern blotting a reactive band in the 3 kb range of PstI-digested genomic DNA was detected. The insertion of these fragments into pUC vectors finally resulted in pc-positive Escherichia coli clones. The gene (encoding a protein with 212 amino acids) was expressed in E. coli with varying deletions at the 5' end. A sequence comparison with other outer membrane proteins of B. burgdorferi indicates a processing of pC that is similar to that of lipoproteins.
我们描述了编码莱姆病螺旋体(Lyme borreliosis)病原体伯氏疏螺旋体(Borrelia burgdorferi)主要免疫显性蛋白的pc基因的克隆与表达。从伯氏疏螺旋体PKo菌株的裂解物中纯化出pC蛋白。对pC蛋白进行胰蛋白酶消化后,将所得的寡肽应用于气相测序仪。由此获得了部分氨基酸序列。推导的寡核苷酸用作杂交探针。经过Southern印迹分析,在经PstI消化的基因组DNA的3 kb范围内检测到一条反应带。将这些片段插入pUC载体最终得到了pc阳性的大肠杆菌克隆。该基因(编码一种含212个氨基酸的蛋白质)在大肠杆菌中表达,5'端有不同程度的缺失。与伯氏疏螺旋体其他外膜蛋白的序列比较表明,pC的加工过程与脂蛋白相似。