title: Human Siglec-5 inhibitory receptor and immunoglobulin A (IgA) have separate binding sites in streptococcal beta protein.
abstract: Sialic acid-binding immunoglobulin-like lectins (Siglecs) are receptors believed to be important for regulation of cellular activation and inflammation. Several pathogenic microbes bind specific Siglecs via sialic acid-containing structures at the microbial surface, interactions that may result in modulation of host responses. Recently, it was shown that the group B Streptococcus (GBS) binds to human Siglec-5 (hSiglec-5), an inhibitory receptor expressed on macrophages and neutrophils, via the IgA-binding surface β protein, providing the first example of a protein/protein interaction between a pathogenic microbe and a Siglec. Here we show that the hSiglec-5-binding part of β resides in the N-terminal half of the protein, which also harbors the previously determined IgA-binding region. We constructed bacterial mutants expressing variants of the β protein with non-overlapping deletions in the N-terminal half of the protein. Using these mutants and recombinant β fragments, we showed that the hSiglec-5-binding site is located in the most N-terminal part of β (B6N region; amino acids 1-152) and that the hSiglec-5- and IgA-binding domains in β are completely separate. We showed with BIAcore(TM) analysis that tandem variants of the hSiglec-5- and IgA-binding domains bind to their respective ligands with high affinity. Finally, we showed that the B6N region, but not the IgA-binding region of β, triggers recruitment of the tyrosine phosphatase SHP-2 to hSiglec-5 in U937 monocytes. Taken together, we have identified and isolated the first microbial non-sialic acid Siglec-binding region that can be used as a tool in studies of the β/hSiglec-5 interaction.
authors.affiliation: Department of Laboratory Medicine, Division of Medical Microbiology, Lund University, Sölvegatan 23, 223 62 Lund, Sweden.
authors.forename: Therése
authors.initials: T
authors.lastname: Nordström
authors.forename: Elin
authors.initials: E
authors.lastname: Movert
authors.forename: Anders I
authors.initials: AI
authors.lastname: Olin
authors.forename: Syed R
authors.initials: SR
authors.lastname: Ali
authors.forename: Victor
authors.initials: V
authors.lastname: Nizet
authors.forename: Ajit
authors.initials: A
authors.lastname: Varki
authors.forename: Thomas
authors.initials: T
authors.lastname: Areschoug
mesh_terms: D000942:Antigens, Bacterial / Q000235:genetics / Q000378:metabolism*; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D015214:Antigens, Differentiation, Myelomonocytic / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D002460:Cell Line; D006801:Humans; D007070:Immunoglobulin A / Q000235:genetics / Q000378:metabolism*; D037102:Lectins / Q000235:genetics / Q000378:metabolism*; D008264:Macrophages / Q000378:metabolism*; D009154:Mutation; D009504:Neutrophils / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013290:Streptococcal Infections / Q000235:genetics / Q000378:metabolism; D013292:Streptococcus agalactiae / Q000235:genetics / Q000378:metabolism
chemical_list: D000942:Antigens, Bacterial; D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D007070:Immunoglobulin A; D037102:Lectins; C114531:SIGLEC5 protein, human; C070010:streptococcal beta antigen