Rao V K, Whitlock J A, Proguske-Fox A
Department of Oral Biology, University of Florida, Gainesville 32610.
J Gen Microbiol. 1993 Mar;139(3):639-50. doi: 10.1099/00221287-139-3-639.
Eikenella corrodens is emerging as an important human pathogen, in both extra-oral and periodontal infections. From a clone bank of Eikenella corrodens chromosomal DNA produced in Escherichia coli JM109, twenty-two clones expressed Eikenella antigens and of these, two expressed functional haemagglutinins. By virtue of different restriction maps and a lack of homology by Southern hybridization, the two cloned fragments encoding the two haemagglutinins have been shown to be distinct. Maxicell analysis revealed that clone 1, carrying plasmid pVKR201, produces three Eikenella proteins, one of 31.5 kDa and two of approximately 14 kDa each. Expression of each of the proteins appears to be under the control of an Eikenella promoter(s). Clone 2, carrying plasmid pVKR301, produces two proteins, one of 93 kDa and the second of 17 kDa. Expression of both of these proteins in E. coli requires the lac promoter in the vector. By preparing a series of subclones and testing each by maxicell analysis and for haemagglutination activity, a functional map of the insert of clone 1 was deduced and the 31.5 kDa polypeptide identified as the haemagglutinin. Using similar methods, the 17 kDa protein was found to be the haemagglutinin of clone 2. The nucleotide sequences of both haemagglutinin genes were determined and are presented. Computer analysis revealed no homology between the two haemagglutinins, and no homology to any previously sequenced proteins. These are the first genes of this genus to be cloned and sequenced.
啮蚀艾肯菌正逐渐成为一种重要的人类病原体，可引发口腔外感染和牙周感染。从在大肠杆菌JM109中构建的啮蚀艾肯菌染色体DNA克隆文库中，有22个克隆表达了啮蚀艾肯菌抗原，其中两个表达了功能性血凝素。通过不同的限制性图谱以及Southern杂交缺乏同源性，已证明编码这两种血凝素的两个克隆片段是不同的。最大细胞分析显示，携带质粒pVKR201的克隆1产生三种啮蚀艾肯菌蛋白，一种为31.5 kDa，另外两种各约为14 kDa。每种蛋白的表达似乎受一个或多个啮蚀艾肯菌启动子的控制。携带质粒pVKR301的克隆2产生两种蛋白，一种为93 kDa，另一种为17 kDa。这两种蛋白在大肠杆菌中的表达需要载体中的lac启动子。通过制备一系列亚克隆，并通过最大细胞分析和血凝活性测试每个亚克隆，推导出了克隆1插入片段的功能图谱，并将31.5 kDa多肽鉴定为血凝素。使用类似方法，发现17 kDa蛋白是克隆2的血凝素。测定并展示了两种血凝素基因（haemagglutinin genes）的核苷酸序列。计算机分析显示这两种血凝素之间没有同源性，也与任何先前测序的蛋白质没有同源性。这些是该属中首次被克隆和测序的基因。
