Structural Characterization of CFA/III and Longus Type IVb Pili from Enterotoxigenic Escherichia coli

Structural and Evolutionary Analyses Show Unique Stabilization Strategies in the Type IV Pili of Clostridium difficile

“…However, unlike FimA, which is a Type IVa pilin and has a continuous β-sheet with relatively short sequences between the strands, PilA1 has a discontinuous β-sheet. The C-terminal β-strand is not the final strand, but instead the α3 helix, the final (fourth) β-strand and the B2 sheet are placed sequentially between the second and third strands; analogous to what is seen in the Type IVb pilins, including TcpA from Vibrio cholorae , CofA from enterotoxigenic Escherichia coli (ETEC) and PilS from S. typhi (Balakrishna et al, 2009; Craig et al, 2003; Kolappan et al, 2012). This leaves a 34- residue loop between the second and fourth strands of the central β-sheet.…”

Crystal Structure of the Minor Pilin CofB, the Initiator of CFA/III Pilus Assembly in Enterotoxigenic Escherichia coli

“…The C-terminal region extends from the "inside" face of the pilin domain that in the major pilins faces into the core of the assembled pilus. The pilin domain has the canonical T4b pilin fold seen in the major pilins, CofA (64) and TcpA (88,89), with an N-terminal ␣-helical backbone, ␣1C, and a 5-stranded anti-parallel ␤-sheet, in which strand ␤5 forms the central ␤-strand. This ␤-sheet lies against ␣1C and two shorter ␣-helices, ␣3 and ␣4, all of which are located on the inside face of the pilin domain (64).…”

“…The ETEC Minor Pilin CofB Is Required for CFA/III Assembly-The ETEC minor pilin CofB is encoded on the cof operon immediately downstream of the gene encoding the major pilin, CofA, the structural subunit for the CFA/III Type IVb pilus (64,79,80). CofB is a 518-amino acid protein with a predicted 5-amino acid signal peptide (Fig.…”

“…Samples were mixed with Laemmli sample buffer (60 mM Tris, pH 6.8, 5% 2-mercaptoethanol, 2% SDS, 10% glycerol, 0.02% bromphenol blue) and boiled for 10 min prior to being loaded onto 15% SDS-polyacrylamide gels. Proteins were transferred onto polyvinylidene difluoride (PVDF) membrane (Bio-Rad) for immunoblotting and detected by rabbit polyclonal antisera raised against N-terminally truncated CofA (64) or CofB (N-terminal peptide 61-74 and C-terminal peptide 403-419; Pacific Immunology). Goat anti-rabbit secondary antibodies conjugated to horseradish peroxidase (Jackson ImmunoResearch) were used to detect primary antibody.…”

Motility and adhesion through type IV pili in Gram-positive bacteria

“…Competence pili [sometimes referred to as type IV filaments rather than type IV pili] in other organisms also lack pilT homologs raising the obvious question of how DNA bound by these appendages is taken up by the cell. However, we note that some bacteria without pilT genes are nevertheless capable of retracting their pili [31,32]. The mechanism for this retraction remains uncharacterized but given the homology between PilB and PilT, it is not unreasonable to propose that an ancestral ATPase was capable of facilitating both extension and retraction, and that some AAA+ ATPases retain the ability to do so.…”