Novel adhesin from Pasteurella multocida that binds to the integrin-binding fibronectin FnIII9-10 repeats.
Phage display screening with fragmented genomic DNA from the animal pathogen Pasteurella multocida has identified a gene encoding a putative fibronectin binding protein (19). Homologues of this gene (PM1665) are found in all other sequenced members of the Pasteurellaceae. Gene PM1665 has been cloned, and the protein has been expressed. Recombinant PM1665 protein binds to both soluble and immobilized fibronectin and is unique in that it interacts with the integrin-binding fibronectin type III (FnIII) repeats FnIII(9-10) and not, as is the case for almost all other fibronectin adhesins, to the N-terminal type I repeats. Surface plasmon resonance analysis revealed a complex binding mechanism with a K(D) (equilibrium dissociation constant) of 150 nM +/- 70 nM. Bioinformatics analysis suggests that the PM1665 protein contains two helix-hairpin-helix (HhH) motifs, and truncation mutation studies have identified the binding site in the protein as a combination of these two HhH motifs in conjunction with a conserved amino acid motif, VNINTA. We have shown that the PM1665 protein is on the cell surface and that binding of P. multocida to fibronectin is almost completely inhibited by anti-PM1665 antiserum. These results support the hypothesis that the PM1665 protein is a member of a new family of fibronectin binding adhesins that are important in the adhesion of P. multocida to fibronectin.
Mullen
Lisa M
LM
Division of Microbial Diseases, Eastman Dental Institute, University College London, 256 Gray's Inn Road, London WC1X 8LD, United Kingdom. l.mullen@eastman.ucl.ac.uk
Nair
Sean P
SP
Ward
John M
JM
Rycroft
Andrew N
AN
Williams
Rachel J
RJ
Robertson
Giles
G
Mordan
Nicky J
NJ
Henderson
Brian
B
D018829:Adhesins, Bacterial / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D003001:Cloning, Molecular; D005353:Fibronectins / Q000378:metabolism*; D015870:Gene Expression; D019077:Helix-Turn-Helix Motifs; D007700:Kinetics; D008565:Membrane Proteins / Q000032:analysis; D016979:Pasteurella multocida / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D019151:Peptide Library; D011485:Protein Binding; D025941:Protein Interaction Mapping; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D020349:Surface Plasmon Resonance
D018829:Adhesins, Bacterial; D005353:Fibronectins; D008565:Membrane Proteins; D019151:Peptide Library; D011994:Recombinant Proteins; C078635:fibronectin-binding proteins, bacterial