Cloning of the Gene Encoding a Major Immunodominant Surface Protein of a Peridontopathogen

本文纯化了牙龈卟啉单胞菌381的主要免疫显性表面蛋白75K蛋白，并测定了其氨基末端氨基酸序列。利用与该序列对应的寡核苷酸探针，从牙龈假单胞菌基因组DNA pUC19文库中鉴定出一个携带4.2千碱基对Bam HI片段的重组质粒克隆。将公羊HI片段转移到噬菌体T7 RNA聚合酶/启动子表达载体系统中，产生一个稍大的77千道尔顿蛋白，这是一种前体形式，对针对75K蛋白的抗体产生免疫反应，表明克隆的DNA片段可能携带75K蛋白的完整基因。基因组南方分析显示，在所有测试的牙龈卟啉杆菌菌株中，每个基因组都有一个75K蛋白基因拷贝，而在其他黑色色素拟杆菌中没有同源基因存在。这些观察结果表明，75K蛋白基因可能是一种有用的特异性DNA探针，用于分类或检测这种生物。

A major immunodominant surface protein (the 75K protein) of Porphyromonas gingivalis 381 has been purified and its amino-terminal amino acid sequence has been determined. Using oligonucleotide probes corresponding to the sequence, we identified a recombinant plasmid clone carrying a single 4.2-kilobase pair Bam HI fragment from pUC19 libraries of P. gingivalis genomic DNA. The Ram HI fragment transferred to the bacteriophage T7 RNA polymerase/promoter expression vector system produced a slightly larger 77-kilodalton protein, a precursor form, immunoreactive to the antibody against the 75K protein, suggesting that the cloned DNA fragment probably carried an entire gene for the 75K protein. Genomic Southern analysis revealed a single copy of the 75K protein gene per genome among all P. gingivalis strains tested, and that no homologous genes are present in other black-pigmented Bacteroides species. These observations suggest that the 75K protein gene may be useful as a specific DNA probe to classify or to detect this organism.