Mapping the binding domains on meningococcal Opa proteins for CEACAM1 and CEA receptors.
The opacity (Opa) proteins of pathogenic Neisseria spp. are adhesins, which play an important role in adhesion and invasion of host cells. Most members of this highly variable family of outer membrane proteins can bind to the human carcinoembryonic antigen-related cell adhesion molecules (CEACAMs). Several studies have identified the Opa-binding region on the CEACAM receptors; however, not much is known about the binding sites on the Opa proteins for the corresponding CEACAM-receptors. The high degree of sequence variation in the surface-exposed loops of Opa proteins raises the question how the binding sites for the CEACAM receptors are conserved. Neisseria meningitidis strain H44/76 possesses four different Opa proteins, of which OpaA and OpaJ bind to CEACAM1, while OpaB and OpaD bind to CEACAM1 and CEA. A sequence motif involved in binding to CEACAM1 was identified by alanine scanning mutagenesis of those amino acid residues conserved within the hypervariable (HV) regions of all four Opa proteins. Hybrid Opa variants with different combinations of HV-1 and HV-2 derived from OpaB and OpaJ showed a reduced binding to CEACAM1 and CEA, indicating that particular combinations of HV-1 and HV-2 are required for the Opa binding capacity. Homologue scanning mutagenesis was used to generate more refined hybrids containing novel combinations of OpaB and OpaJ sequences within HV-1 and HV-2. They could be used to identify residues determining the specificity for CEA binding. The combined results obtained with mutants and hybrids strongly suggest the existence of a conserved binding site for CEACAM receptors by the interaction of HV-1 and HV-2 regions.
de Jonge
Marien I
MI
Laboratory of Vaccine Research, Netherlands Vaccine Institute, Bilthoven, the Netherlands.
Hamstra
Hendrik Jan
HJ
van Alphen
Loek
L
Dankert
Jacob
J
van der Ley
Peter
P
D000409:Alanine / Q000235:genetics; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D000943:Antigens, Differentiation / Q000235:genetics / Q000378:metabolism*; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D015815:Cell Adhesion Molecules; D002462:Cell Membrane / Q000378:metabolism; D006367:HeLa Cells / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009154:Mutation; D009345:Neisseria meningitidis / Q000378:metabolism / Q000472:pathogenicity; D010449:Peptide Mapping / Q000379:methods; D011956:Receptors, Cell Surface; D011975:Receptors, Mitogen / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
D015703:Antigens, CD; D000943:Antigens, Differentiation; D001425:Bacterial Outer Membrane Proteins; C069847:CD66 antigens; D002352:Carrier Proteins; D015815:Cell Adhesion Molecules; D011956:Receptors, Cell Surface; D011975:Receptors, Mitogen; D011994:Recombinant Proteins; C065686:carcinoembryonic antigen binding protein, human; C090348:Opa protein, Neisseria; D000409:Alanine