1887

Abstract

is the causative agent of epidemic meningococcal meningitis and septicaemia. Type IV pili are surface organelles that mediate a variety of functions, including adhesion, twitching motility, and competence for DNA binding and uptake in transformation. The secretin PilQ is required for type IV pilus expression at the cell surface, and forms a dodecameric cage-like macromolecular complex in the meningococcal outer membrane. PilQ-null mutants are devoid of surface pili, and prevailing evidence suggests that the PilQ complex facilitates extrusion and retraction of type IV pili across the outer membrane. Defining the orientation of the meningococcal PilQ complex in the membrane is a prerequisite for understanding the structure–function relationships of this important protein in pilus biology. In order to begin to define the topology of the PilQ complex in the outer membrane, polyhistidine insertions in N- and C-terminal regions of PilQ were constructed, and their subcellular locations examined. Notably, the insertion epitopes at residues 205 and 678 were located within the periplasm, whereas residue 656 was exposed at the outer surface of the outer membrane. Using electron microscopy with Ni-NTA gold labelling, it was demonstrated that the insertion at residue 205 within the N-terminus mapped to a site on the arm-like features of the 3D structure of the PilQ multimer. Interestingly, mutation of the same region gave rise to an increase in vancomycin permeability through the PilQ complex. The results yield novel information on the PilQ N-terminal location and function in the periplasm, and reveal a complex organization of the membrane-spanning secretin .

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2006-12-01
2019-10-13
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A table showing predictions of the C-terminal βstrands for PilQ from strain M1080 is available hereas an Acrobat PDF file.

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A table showing predictions of the C-terminal βstrands for PilQ from strain M1080 is available hereas an Acrobat PDF file.

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Secretin alignment. clustalx (Thompson , 1997) alignment of the C-terminal parts of XcpQ (GenBank accession no. AAG06493) and strain M1080 PilQ (CAD91899), starting with amino acids 322 and 463, respectively. Predicted β strands for XcpQ (Bitter , 1998) and PilQ (Gromiha , 1997) are highlighted. The positions of the His-tag insertions in PilQ are marked with arrows indicating the amino acids which are followed by the tags. Identical residues: 23 %; similar residues 41 % (in the alignment shown). The His-tag insertion positions for Mc-656 and Mc-678 are predicted to be located in neighbouring loops.

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