Identification of class II major histocompatibility complex and T cell receptor binding sites in the superantigen toxic shock syndrome toxin 1.

Abstract

SummarySuperantigens, in association with class II major histocompatibility complex (MHC) molecules, activate T cells bearing particular/~ chain variable domains of the T cell receptor (TCR). Unlike conventional peptide antigens, superantigens bind as intact proteins to TCR and MHC molecules outside their peptide binding sites. To characterize these interactions at the molecular level, random point mutations were generated in the gene encoding toxic shock syndrome toxin 1, a bacterial superantigen associated with toxic shock syndrome. Functionally impaired mutants were identified based on their lack of murine and human T cell stimulatory activities, and experiments analyzing binding to human histocompatibility leukocyte antigen-DR molecules differentiated residues involved in MHC from TCR binding. The results showed that the great majority of mutations are clustered in two distinct regions of the toxic shock syndrome toxin 1 molecule. The class II MHC binding site is located in the hydrophobic region of the NH2-terminal domain, and the TCR binding site is primarily in the major central groove of the COOH-terminal domain. These studies provide insight into the interactions necessary for superantigen-mediated disease in humans.

Functional Analysis of the TCR Binding Domain of Toxic Shock Syndrome Toxin-1 Predicts Further Diversity in MHC Class II/Superantigen/TCR Ternary Complexes

“…Mitogenicity assays typically exhibited a clear dose response, where wild-type mitogenic capacity was achieved with SAG concentrations at 50 pg/ml, similar to previous data (30,31,39). For some protein preparations, the highest dose tested (5 g/ml) occasionally had reduced activity or failed to stimulate proliferation.…”

“…Alanine substitution was chosen because this technique removes the targeted amino acid side chain while minimizing steric or electrostatic constrains on the tertiary structure of the protein (37 (30), which is located on the N-terminal ␣ helix, as well as E132K and Q136A, located along the central helix (31). A control MHC class II binding mutant was also created (G31S/S32P) that lacks significant proliferative activity (30), and these residues are known to be located at the interface with the MHC class II molecule HLA-DR1 (16). The Ser 32 mutation is believed to account for most of the impaired interaction with MHC class II (30), although other data suggest that Gly 31 is also critical for MHC class II binding (38).…”

“…A control MHC class II binding mutant was also created (G31S/S32P) that lacks significant proliferative activity (30), and these residues are known to be located at the interface with the MHC class II molecule HLA-DR1 (16). The Ser 32 mutation is believed to account for most of the impaired interaction with MHC class II (30), although other data suggest that Gly 31 is also critical for MHC class II binding (38). All the mutant genes (Table I) were constructed using plasmid pCE107 as a template, and mutated proteins were expressed and purified from S. aureus RN4220.…”

The Staphylococcal Enterotoxins

“…In general, there is a distinct functional separation of the SE N-terminal and Cterminal domains, whereby one domain is responsible for SE interaction with TCR, while the other domain mediates SE-MHC II interaction (Kappler et al, 1992;Swaminathan et al, 1992;Soos et al, 1993;Hurley et al, 1995;Mahana et al, 1995;Li et al, 1998a, b), but die specificities vary between different members of the SE family. The SE-TCR interaction appears to play a pivotal role in the superantigenic activity of SEs.…”

Analysis of the interaction between the bacterial superantigen streptococcal pyrogenic exotoxin A (SpeA) and the human T‐cell receptor

“…Each superantigenic toxin examined seems to interact with the TCR in a unique fashion. Staphylococcal TSST-1 residues that contact the TCR are found primarily in the central groove of the C-terminal domain (Hurley et al, 1995). Conversely, SEB and SEC1 both bind the TCR via N-terminal toxin residues (Hoffman et al, 1994;Kappler et al, 1992), and SEA requires N-and C-terminal toxin residues to form TCR contact sites (Mollick et al, 1993).…”