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. 2017 Feb 14;292(14):5724–5735. doi: 10.1074/jbc.M117.776724

FIGURE 3.

FIGURE 3.

Effect of Lys129 mutation on Kgp expression and activity. Western blotting analysis of parental strain P. gingivalis W83 and mutants K129A, K129E, and K129R is shown. Late exponential/early stationary bacterial cultures (BC) were separated by centrifugation into cell-free growth medium (GM) and cell pellet. The latter was washed, giving rise to the whole cell (WC) fraction, which was further fractionated into the soluble intracellular protein fraction (CP + PP) and the cell envelope fracion (CE) by sonication and ultracentrifugation. A and B, all fractions were standardized to the initial volume of the culture subjected to centrifugation and analyzed by Western blotting to detect Kgp forms (A) and Rgps (control) (B). C and D, Kgp (C) and Rgp (D) gingipain activities determined in whole cultures, cell-free growth medium, and washed cells using specific substrates. The whole-culture activity of the wild-type strain was arbitrarily taken as 100%.

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