Role of sialic acid-containing glycans of matrix metalloproteinase-9 (MMP-9) in the interaction between MMP-9 and staphylococcal superantigen-like protein 5.

Staphylococcal superantigen-like proteins (SSL) show no superantigenic activity but have recently been considered to act as immune suppressors. It was previously reported that SSL5 bound to P-selectin glycoprotein ligand-1 (PSGL-1) and matrix metalloproteinase (MMP)-9, leading to inhibition of leukocyte adhesion and invasion. These interactions were suggested to depend on sialic acid-containing glycans of MMP-9, but the roles of sialic acids in the interaction between SSL5 and MMP-9 are still controversial. In the present study, we prepared recombinant glutathione S-transferase-tagged SSL5 (GST-SSL5) and analyzed its binding capacity to MMP-9 by pull-down assay after various modifications of its carbohydrate moieties. We observed that GST-SSL5 specifically bound to MMP-9 from a human monocytic leukemia cell line (THP-1 cells) and inhibited its enzymatic activity in a concentration-dependent manner. After MMP-9 was treated with neuraminidase, its binding activity towards GST-SSL5 was markedly decreased. Furthermore, recombinant MMP-9 produced by sialic acid-deficient Lec2 mutant cells showed much lower affinity for SSL5 than that produced by wild-type CHO-K1 cells. Treatment of MMP-9 with PNGase F to remove N-glycan resulted in no significant change in the GST-SSL5/MMP-9 interaction. In contrast, the binding of GST-SSL5 to MMP-9 secreted from THP-1 cells cultured in the presence of an inhibitor for the biosynthesis of O-glycan (benzyl-GalNAc) was weaker than the binding of GST-SSL5 to MMP-9 secreted from untreated cells. These results strongly suggest the importance of the sialic acid-containing O-glycans of MMP-9 for the interaction of MMP-9 with GST-SSL5.