Porphyromonas gingivalis uses a type IX secretion system (T9SS) to deliver more than 30 proteins to the bacterial surface using a conserved C-terminal domain (CTD) as an outer membrane translocation signal. On the surface, the CTD is cleaved and an anionic lipopolysaccharide (A-PLS) is attached by PorU sortase. Among T9SS cargo proteins are cysteine proteases, gingipains, which are secreted as inactive zymogens requiring removal of an inhibiting N-terminal prodomain (PD) for activation. Here, we have shown that the gingipain proRgpB isolated from the periplasm of a T9SS-deficient P. gingivalis strain was stable and did not undergo autocatalytic activation. Addition of purified, active RgpA or RgpB, but not Lys-specific Kgp, efficiently cleaved the PD of proRgpB but catalytic activity remained inhibited because of inhibition of the catalytic domain in trans by the PD. In contrast, active RgpB was generated from the zymogen, although at a slow rate, by gingipain-null P. gingivalis lysate or intact bacterial cell suspension. This activation was dependent on the presence of the PorU sortase. Interestingly, maturation of proRgpB with the catalytic cysteine residues mutated to Ala expressed in the ΔRgpA mutant strain was indistinguishable from that in the parental strain. Cumulatively, this suggests that PorU not only has sortase activity but is also engaged in activation of gingipain zymogens on the bacterial cell surface.

中文翻译：

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牙龈卟啉单胞菌的T9SS分泌的蛋白水解酶处理和激活。

牙龈卟啉单胞菌使用IX型分泌系统（T9SS），利用保守的C末端结构域（CTD）作为外膜易位信号，将30多种蛋白质传递到细菌表面。在表面上，CTD被切割，阴离子脂多糖（A-PLS）通过PorU分选酶连接。在T9SS货物蛋白中，半胱氨酸蛋白酶，牙龈蛋白酶被分泌为非活性酶原，需要去除抑制性N端前结构域（PD）才能激活。在这里，我们已经表明，从T9SS缺陷型牙龈卟啉单胞菌菌株的周质中分离出的gingipain proRgpB是稳定的，并且未经历自动催化活化。加入纯化的活性RgpA或RgpB，但不添加Lys特异性Kgp，高效地切割了proRgpB的PD，但是由于PD对反式催化结构域的抑制，催化活性仍然受到抑制。相比之下，尽管是缓慢的，但通过gingipain-null gingivalis裂解物或完整的细菌细胞悬液从酶原中产生了活性RgpB。该激活取决于PorU分选酶的存在。有趣的是，proRgpB的成熟与在ΔRgpA突变株中表达的突变为Ala的催化半胱氨酸残基没有区别。累积地，这表明PorU不仅具有分选酶活性，而且还参与细菌细胞表面上的姜黄素酶原的活化。牙龈溶解物或完整的细菌细胞悬液。该激活取决于PorU分选酶的存在。有趣的是，proRgpB的成熟与在ΔRgpA突变株中表达的突变为Ala的催化半胱氨酸残基没有区别。累积地，这表明PorU不仅具有分选酶活性，而且还参与细菌细胞表面上的姜黄素酶原的活化。牙龈溶解物或完整的细菌细胞悬液。该激活取决于PorU分选酶的存在。有趣的是，proRgpB的成熟与在ΔRgpA突变株中表达的突变为Ala的催化半胱氨酸残基没有区别。累积地，这表明PorU不仅具有分选酶活性，而且还参与细菌细胞表面上的姜黄素酶原的活化。