Human Siglec-5 Inhibitory Receptor and Immunoglobulin A (IgA) Have Separate Binding Sites in Streptococcal β 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 BIAcoreTM 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.

The Combined Role of Galactose-Deficient IgA1 and Streptococcal IgA–Binding M Protein in Inducing IL-6 and C3 Secretion from Human Mesangial Cells: Implications for IgA Nephropathy

“…The IgA-binding M4 and non-IgA-binding M4Δ451 were diluted in 10 mM sodium acetate (pH 4) (Merck, Darmstadt, Germany) and immobilized via amine coupling on separate CM5 sensor chip chambers (GE Healthcare, Uppsala, Sweden) at moderate response levels (1500 response units) as previously described (44). A flow chamber subjected to the immobilization conditions but without addition of protein was used as a control (blank) for each experiment.…”

Finding the Smoking Gun: Protein Tyrosine Phosphatases as Tools and Targets of Unicellular Microorganisms and Viruses

“…Several serotypes of the group B Streptococci (serotypes Ia, Ib, II and V) are known to produce the Streptococcus β protein [13], which binds SHP-2 directly. SHP-2 binds to the Nterminal part of Streptococcus β protein (AA 1-152, called B6N region); this region is sufficient to trigger recruitment of SHP-2 to Siglec-5 (tested using the human monocyte cell line U937) [14]. Exact role of SHP-2 in the Streptococcus β protein signaling is poorly understood; use of the isogenic β-negative group B Streptococcus mutant revealed that the Streptococcus β protein interaction with host Siglec-5 recruits SHP-2 to Siglec-5 in U937 cells, inhibits phagocytosis of group B Streptococci by U937 cells, inhibits the IL-8 secretion of primary human neutrophils, and suppresses also the oxidative burst of primary human neutrophils and formation of neutrophil extracellular traps in response to the group B Streptococci [14].…”

“…Selected inhibitors of non-metazoan PTPs:(12) benzanilide scaffold-based compound,(13) isoxazole carboxylic acid isostere 433 Da analogue,(14) brunsvicamide B, (15) brunsvicamide C,(16) iodoacetamide, (17) N-ethylmaleimide,(18)(19) isothiazolidinone derivatives, (20) nitrophenylphosphate derivative, (21) bidentate α-ketoacid-based compound, (22) salicylic acid derivative, (23) homotyrosine-based selenic acid.…”

Perinatal Group B Streptococcal Infections: Virulence Factors, Immunity, and Prevention Strategies

“…GBS MVs contained multiple virulence factors, including: 1) HylB, 2) CAMP factor (Christine, Atkins, Munch-Peterson factor [85]), a secreted pore-forming protein [86] that may amplify [87], but is not essential for GBS virulence [88], 3) IgA binding protein, with the ability to bind human IgA [89] for host immune evasion [90] and 4) multiple enzymes that may regulate ECM degradation [84]. Intra-amniotic administration of GBS MVs in pregnant mice caused significant damage to choriodecidual tissues and stimulated leukocytic infiltration and inflammation, leading to membrane weakening [84].…”