Aduse-Opoku J, Slaney J M, Rangarajan M, Muir J, Young K A, Curtis M A
Department of Oral Microbiology, St. Bartholomew's and the Royal London School of Medicine and Dentistry, Queen Mary and Westfield College, England.
J Bacteriol. 1997 Aug;179(15):4778-88. doi: 10.1128/jb.179.15.4778-4788.1997.
The prpR1 gene of Porphyromonas gingivalis W50 encodes the polyprotein precursor (PrpRI) of an extracellular arginine-specific protease. PrpRI is organized into four distinct domains (pro, alpha, beta, and gamma) and is processed to a heterodimeric protease (RI) which comprises the alpha and beta components in a noncovalent association. The alpha component contains the protease active site, whereas the beta component appears to have a role in adherence and hemagglutination processes. DNA sequences homologous to the coding region for the RI beta component are present at multiple loci on the P. gingivalis chromosome and may represent a family of related genes. In this report, we describe the cloning, sequence analysis, and characterization of one of these homologous loci isolated in plasmid pJM7. The 6,041-bp P. gingivalis DNA fragment in pJM7 contains a major open reading frame of 3,291 bp with coding potential for a protein with an Mr 118,700. An internal region of the deduced sequence (V304 to N768) shows 98% identity to the beta domain of PrpRI, and the recombinant product of pJM7 is immunoreactive with an antibody specific to the RI beta component. The N terminus of the deduced sequence has regional similarity to TonB-linked receptors which are frequently involved in periplasmic translocation of hemin, iron, colicins, or vitamin B12 in other bacteria. We have therefore designated this gene tla (TonB-linked adhesin). In contrast to the parent strain, an isogenic mutant of P. gingivalis W50 in which the tla was insertionally inactivated was unable to grow in medium containing low concentrations of hemin (<2.5 mg liter(-1)), and hemin-depleted cells of this mutant failed to respond to hemin in an agar diffusion plate assay. These data suggest a role for this gene product in hemin acquisition and utilization. Furthermore, the mutant produced significantly less arginine- and lysine-specific protease activities than the parent strain, indicating that there may be a regulatory relationship between tla and other members of this gene family.
牙龈卟啉单胞菌W50的prpR1基因编码一种细胞外精氨酸特异性蛋白酶的多蛋白前体（PrpRI）。PrpRI由四个不同的结构域（前导区、α区、β区和γ区）组成，并被加工成一种异二聚体蛋白酶（RI），该蛋白酶由α和β组分通过非共价结合构成。α组分含有蛋白酶活性位点，而β组分似乎在黏附和血凝过程中起作用。与RI β组分编码区同源的DNA序列存在于牙龈卟啉单胞菌染色体的多个位点，可能代表一个相关基因家族。在本报告中，我们描述了从质粒pJM7中分离出的这些同源位点之一的克隆、序列分析和特性鉴定。pJM7中6041 bp的牙龈卟啉单胞菌DNA片段包含一个3291 bp的主要开放阅读框，编码一种Mr为118700的蛋白质。推导序列的内部区域（V304至N768）与PrpRI的β结构域有98%的同一性，pJM7的重组产物与RI β组分特异性抗体发生免疫反应。推导序列的N端与TonB连接受体有区域相似性，TonB连接受体在其他细菌中经常参与血红素、铁、大肠杆菌素或维生素B12的周质转运。因此，我们将该基因命名为tla（TonB连接黏附素）。与亲本菌株相比，牙龈卟啉单胞菌W50的一个等位基因突变体，其中tla被插入失活，在含有低浓度血红素（<2.5 mg L⁻¹）的培养基中无法生长，并且该突变体的血红素耗尽细胞在琼脂扩散平板试验中对血红素无反应。这些数据表明该基因产物在血红素获取和利用中起作用。此外，该突变体产生的精氨酸和赖氨酸特异性蛋白酶活性明显低于亲本菌株，表明tla与该基因家族的其他成员之间可能存在调控关系。
