The Gram-negative anaerobe, Porphyromonas gingivalis, is known to be a pathogen associated with chronic periodontitis. P. gingivalis possesses virulence factors such as fimbriae and gingipain proteinases. Fimbrial proteins are secreted to the cell surface as lipoproteins. In contrast, gingipain proteinases are secreted into the bacterial cell surface via the type IX secretion system (T9SS). The transport mechanisms of lipoproteins and T9SS cargo proteins are entirely different and remain unknown. Therefore, using the Tet-on system developed for the genus Bacteroides, we newly created a conditional gene expression system in P. gingivalis. We succeeded in establishing conditional expression of nanoluciferase and its derivatives for lipoprotein export, of FimA for a representative of lipoprotein export, and of T9SS cargo proteins such as Hbp35 and PorA for representatives of type 9 protein export. Using this system, we showed that the lipoprotein export signal, which has recently been found in other species in the phylum Bacteroidota, is also functional in FimA, and that a proton motive force inhibitor can affect type 9 protein export. Collectively, our conditional protein expression method is useful for screening inhibitors of virulence factors, and may be used to investigate the role of proteins essential to bacterial survival in vivo.

中文翻译：

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牙龈卟啉单胞菌条件基因表达系统，用于研究脂蛋白和 T9SS 货物蛋白的分泌机制

已知革兰氏阴性厌氧菌牙龈卟啉单胞菌是与慢性牙周炎相关的病原体。牙龈卟啉单胞菌具有菌毛和牙龈蛋白酶等毒力因子。菌毛蛋白作为脂蛋白分泌到细胞表面。相反，牙龈蛋白酶通过 IX 型分泌系统 (T9SS) 分泌到细菌细胞表面。脂蛋白和 T9SS 货物蛋白的转运机制完全不同，目前仍不清楚。因此，利用针对拟杆菌属开发的Tet-on系统，我们在牙龈卟啉单胞菌中新创建了条件基因表达系统。我们成功地建立了用于脂蛋白输出的纳米荧光素酶及其衍生物的条件表达、用于脂蛋白输出的代表的 FimA 以及用于代表 9 型蛋白输出的 T9SS 货物蛋白（如 Hbp35 和 PorA）的条件表达。使用该系统，我们表明最近在拟杆菌门其他物种中发现的脂蛋白输出信号在 FimA 中也有功能，并且质子动力抑制剂可以影响 9 型蛋白输出。总的来说，我们的条件蛋白表达方法可用于筛选毒力因子抑制剂，并可用于研究细菌体内生存所必需的蛋白质的作用。