Okamoto K, Nakayama K, Kadowaki T, Abe N, Ratnayake D B, Yamamoto K
Department of Pharmacology, Kyushu University Faculty of Dentistry, Higashi-ku, Fukuoka 812-8582, Japan.
J Biol Chem. 1998 Aug 14;273(33):21225-31. doi: 10.1074/jbc.273.33.21225.
The oral anaerobic bacterium Porphyromonas gingivalis, a major pathogen of advanced adult periodontitis, produces a novel class of cysteine proteinases in both cell-associated and secretory forms. A lysine-specific cysteine proteinase (Lys-gingipain, KGP), as well as an arginine-specific cysteine proteinase (Arg-gingipain), is a major trypsin-like proteinase of the organism. Recent studies indicate that the secreted KGP is implicated in the destruction of periodontal tissue and the disruption of host defense mechanisms. In this study, we have constructed a KGP-deficient mutant to determine whether the cell-associated KGP is important for pathophysiology of the organism. Although the mutant retained the strong ability to disrupt the bactericidal activity of polymorphonuclear leukocytes, its hemagglutination activity was reduced to about one-half that observed with the wild-type strain. More important, the mutant did not form black-pigmented colonies on blood agar plates, indicating the defect of hemoglobin adsorption and heme accumulation. Immunoblot analysis showed that the expression of a 19-kDa hemoglobin receptor protein, which is thought to be responsible for hemoglobin binding by the organism, was greatly retarded in this mutant. The mutant also showed a marked decrease in the ability to degrade fibrinogen. These results suggest the possible involvement of KGP in the hemoglobin binding and heme accumulation of the organism and in the bleeding tendency in periodontal pockets.
口腔厌氧菌牙龈卟啉单胞菌是成人晚期牙周炎的主要病原体，它能产生一类新的细胞相关型和分泌型半胱氨酸蛋白酶。赖氨酸特异性半胱氨酸蛋白酶（Lys-牙龈蛋白酶，KGP）以及精氨酸特异性半胱氨酸蛋白酶（Arg-牙龈蛋白酶）是该菌主要的类胰蛋白酶。最近的研究表明，分泌型KGP与牙周组织的破坏以及宿主防御机制的破坏有关。在本研究中，我们构建了一个KGP缺陷型突变体，以确定细胞相关型KGP对该菌病理生理学是否重要。尽管该突变体仍保留破坏多形核白细胞杀菌活性的强大能力，但其血凝活性降至野生型菌株的约二分之一。更重要的是，该突变体在血琼脂平板上不形成黑色色素菌落，表明其血红蛋白吸附和血红素积累存在缺陷。免疫印迹分析表明，一种被认为负责该菌血红蛋白结合的19 kDa血红蛋白受体蛋白在该突变体中的表达大大延迟。该突变体降解纤维蛋白原的能力也显著降低。这些结果表明，KGP可能参与了该菌的血红蛋白结合和血红素积累以及牙周袋的出血倾向。
