Analysis of major outer membrane proteins from periodontopathogen and relationship between these proteins and clinical symptom in periodontitis

1.不同菌株的主要外膜蛋白分析：各菌株的主要外膜蛋白均为raga和Pgm6/7，分布规律相似。2.培养环境因素对牙龈假单胞菌主要外膜蛋白表达的影响当考察多种因素对主要外膜蛋白表达的影响时，温度是最重要的因素。随着温度的升高，RAGA、RagB和牙龈痛以及菌毛和短菌毛的表达减少。然后，我们采用连续培养，其中可能的培养环境调节。随着生长速度的增加，菌毛和短菌毛的产量增加，而牙周疼痛的数量和活性下降。在营养不良的条件下，菌毛和短菌毛减少。当气氛改变时，…从厌氧状态到好氧状态，牙周疼痛活性降低。3.利用基因缺失突变体对主要外膜蛋白进行鉴定。外膜蛋白Pgm6/7与大肠杆菌OmpA蛋白同源，存在于外膜形成的异源三聚体中。由于Pgm6/7基因缺失突变体在电子显微镜下观察到大量的囊泡状结构，Pgm6/7可能参与了外膜完整性的稳定。由于raga和ragB缺失突变体的构建已经完成，我们正在对突变体进行鉴定。4.牙周炎患者血清对牙周炎主要外膜蛋白的免疫反应。半数以上的血清对牙龈假单胞菌细胞成分有较强的免疫反应，尤其是外膜蛋白RagB和牙龈痛。临床症状如敲击、根尖周半透明与牙周炎的免疫反应性有关。较少

1.Analysis of major outer membrane proteins among various P.gingivalis strains.Major outer membrane proteins, such as RagA and Pgm6/7,were found in all strains tested in a similar distribution pattern. However, mobility and reaction of RagB against specific antibody was different among strains, indicating RagB heterogeneity.2.Effect of culture environment factors on the expression of P.gingivalis major outer membrane proteinsWhen effect of several factors was examined on the expression of major outer membrane proteins, temperature was the most important factor. As temperature increased, expression of RagA,RagB and gingipains as well as fimbriae and short fimbriae decreased.Then, we used continuous culture in which regulation of culture environment was possible. With increase of growth rate, production of fimbriae and short fimbriae increased, while amount and activity of gingipains decreased. In a nutrient-poor medium, fimbriae and short fimbriae decreased. When atmosphere was changed … More from anaerobic to aerobic condition, activity of gingipains decreased. Protein spots whose amount increased under oxidative stress was observed by 2-dimensional gel electrophoresis, and those were identified by mass spectrometry3.Characterization of major outer membrane proteins using gene-deletion mutants.Major outer membrane proteins Pgm6/7, which were homologous to OmpA protein from Escherichia coli, existed in outer membrane forming heterotrimers. Since numerous vesicle-like structures were observed in the case of Pgm6/7 gene-deletion mutants by electron microscopy, Pgm6/7 might be involved in stabilization of outer membrane integrity. Since construction of RagA and RagB-deletion mutants have been finished, we are characterizing the mutants.4.Immunological reactions of sera from patients with periodontitis to P.gingivalis major outer membrane proteins.To examine immunoreactive antigens in P.gingivalis, sera from patients with periapical periodontitis were used. More than half of the sera showed strong immunological reaction to P.gingivalis cell components, especially, outer membrane proteins RagB and gingipains. Clinical symptoms such as percussion and periapical translucency were related to immnoreactivity to P.gingivalis. Less