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Staphylococcus aureus is a prevalent and significant human pathogen. Among the repertoire of virulence factors produced by this bacterium are the 14 staphylococcal superantigen-like (SSL) proteins. SSL protein 4 (SSL4) is one member of this family and contains a highly conserved carbohydrate binding site also found in SSL2, SSL3, SSL5, SSL6, and SSL11. Recombinant SSL4(t), comprising amino acids 109 to 309 of Newman strain SSL4 (SSL4-Newman), has been shown to bind and be internalized by human granulocytes and macrophages in a sialic-acid (Sia)-dependent manner. SSL4(t) can compete with itself for cell binding, indicating that binding is target specific. A 2.5-Å-resolution crystal structure of SSL4(t) complexed with sialyl Lewis X (sLe(x)) [sLe(x)-Neu5Acα2-3Galβ1-4(Fucα1-3)GlcNAc] revealed a similar binding site to SSL5 and SSL11. These data, along with data on SSL4(t) binding to a glycan array and biosensor analysis of sLe(x) and sialyllactosamine (sLacNac) binding are compared with those for SSL11. Although these proteins show great similarity in their carbohydrate binding sites, with a root mean square (RMS) difference between main chain atom positions of only 0.34 Å, these proteins differ in detail in their affinity for sLe(x) and sLacNac, as well as their glycan preference. Together with cell binding data, this shows how S. aureus produces multiple related proteins that target myeloid cells through specific sialyllactosamine-containing glycoproteins.

Staphylococcus aureus is an opportunistic pathogen that produces many virulence factors. Two major families of which are the staphylococcal superantigens (SAgs) and the Staphylococcal Superantigen-Like (SSL) exoproteins. The former are immunomodulatory toxins that induce a Vβ-specific activation of T cells, while the latter are immune evasion molecules that interfere with a wide range of innate immune defences. The superantigenic properties of Staphylococcal enterotoxin-like X (SElX) have recently been established. We now reveal that SElX also possesses functional characteristics of the SSLs. A region of SElX displays high homology to the sialyl-lactosamine (sLacNac)-specific binding site present in a sub-family of SSLs. By analysing the interaction of SElX with sLacNac-containing glycans we show that SElX has an equivalent specificity and host cell binding range to the SSLs. Mutation of key amino acids in this conserved region affects the ability of SElX to bind to cells of myeloid origin and significantly reduces its ability to protect S. aureus from destruction in a whole blood killing (WBK) assay. Like the SSLs, SElX is up-regulated early during infection and is under the control of the S. aureus exotoxin expression (Sae) two component gene regulatory system. Additionally, the structure of SElX in complex with the sLacNac-containing tetrasaccharide sialyl Lewis X (sLeX) reveals that SElX is a unique single-domain SAg. In summary, SElX is an ‘SSL-like’ SAg. [ABSTRACT FROM AUTHOR]