Joe A, Yamamoto A, McBride B C
Department of Microbiology, University of British Columbia, Vancouver, Canada.
Infect Immun. 1993 Aug;61(8):3294-303. doi: 10.1128/iai.61.8.3294-3303.1993.
The cloning of genes encoding putative cell surface antigens of Porphyromonas gingivalis ATCC 33277 has been reported previously (B. C. McBride, A. Joe, and U. Singh, Arch. Oral Biol. 55:59S-68S, 1990). This study characterizes the recombinant protein rPgAg1, which is highly expressed in clone BA3, and the corresponding 51-kDa native antigen PgAg1. Cellular localization studies with monospecific antibodies to rPgAg1 in a Western immunoblot assay of a P. gingivalis membrane fraction and immunogold labeling of intact P. gingivalis cells confirmed the cell surface location of the native PgAg1 molecule. The pgag1 gene was found to be present in all four strains of P. gingivalis examined, and the gene product was expressed. Highly homologous DNA sequences and immunologically related proteins, however, were not detected in related species in the group formerly known as black-pigmented Bacteroides. This suggests that PgAg1 is specific to P. gingivalis and is highly conserved within this species. A protein data base search with the NH2-terminal amino acid sequence of rPgAg1 did not identify any significantly similar protein sequences. The high level of expression of rPgAg1 was not dependent on the insertional orientation of the cloned fragment. It therefore appears that a P. gingivalis promoter is present which is well recognized by the transcriptional apparatus of the Escherichia coli cloning host. The promoter element and structural gene for a specific cell surface antigen of P. gingivalis have been cloned.
先前已有报道克隆牙龈卟啉单胞菌ATCC 33277假定细胞表面抗原的编码基因（B.C.麦克布赖德、A.乔和U.辛格，《口腔生物学文献》55:59S - 68S，1990年）。本研究对在克隆BA3中高表达的重组蛋白rPgAg1以及相应的51 kDa天然抗原PgAg1进行了表征。在牙龈卟啉单胞菌膜组分的蛋白质免疫印迹分析中，用针对rPgAg1的单特异性抗体进行细胞定位研究，以及对完整的牙龈卟啉单胞菌细胞进行免疫金标记，证实了天然PgAg1分子位于细胞表面。发现pgag1基因存在于所检测的所有四株牙龈卟啉单胞菌中，且该基因产物得到表达。然而，在以前称为产黑色素拟杆菌属的相关菌种中未检测到高度同源的DNA序列和免疫相关蛋白。这表明PgAg1是牙龈卟啉单胞菌特有的，且在该菌种内高度保守。用rPgAg1的NH2末端氨基酸序列进行蛋白质数据库搜索，未发现任何显著相似的蛋白质序列。rPgAg1的高表达水平不依赖于克隆片段的插入方向。因此，似乎存在一个牙龈卟啉单胞菌启动子，它能被大肠杆菌克隆宿主的转录装置很好地识别。牙龈卟啉单胞菌一种特定细胞表面抗原的启动子元件和结构基因已被克隆。
