(A) fimA-null mutants constructed to study the impact of truncating the C-terminus of Mfa2 on the length of the Mfa1 pili. 1: fimA-null of P. gingivalis 33277, 2: mfa2-null of the mutant 1, 3: Mutant 2 complemented with mfa2+ on a plasmid, 4: Mutant 2 complemented with mfa2[C29A] on a plasmid, and 5-7: Mutant 2 complemented C-terminal truncated of mfa2 (residues 1-303, 1-314 or 1-321).
(B) Expression of Mfa2 and Mfa1 in mutants 1-7 detected by antibodies.
(C) Cell surface presence of Mfa2 analyzed by dot blots. Cell surface protein HBP35 was used as a control.
(D) Average length (nm) of the Mfa1 pili in mutants 1-7. Error bars represent the standard deviation. * P<0.001.
(E) Representative EM images of mutants 1-7 (pili are marked by arrows).
(F) Mfa1 and Mfa2 interactions in mutants analyzed by western blots (Mfa2-C29A does not interact with Mfa1 since it cannot reach the OM). Asterisk, non-specific bands.