A mutation at glycine residue 31 of toxic shock syndrome toxin-1 defines a functional site critical for major histocompatibility complex class II binding and superantigenic activity.
Random mutagenesis with hydroxylamine was done on toxic shock syndrome toxin-1 (TSST-1) to identify the amino acid residues critical for binding to major histocompatibility complex (MHC) class II molecules of human monocytes. A double mutant with amino acid substitutions of glycine 31-->arginine and aspartic acid 184-->asparagine (G31R.D184N) demonstrated markedly reduced binding to human monocytes and induction of mitogenesis or cytokine secretion. Site-directed mutagenesis revealed that G31R, but not D184N, was at least 4 orders of magnitude less active than wild type recombinant (r) TSST-1 in these biologic activities and did not induce lethal shock in mice. The global structure of G31R remained highly similar to wild type rTSST-1 as evidenced by circular dichroism spectroscopy and binding to anti-TSST-1 polyclonal and monoclonal antibodies. These studies identified TSST-1 residue 31 as critical for binding to MHC class II molecules and for the consequent superantigenic and lethal properties of TSST-1.
Kum
W W
WW
Department of Medicine, University of British Columbia, Canada.
Wood
J A
JA
Chow
A W
AW
D000818:Animals; D000907:Antibodies, Bacterial / Q000276:immunology; D001224:Aspartic Acid / Q000235:genetics; D001427:Bacterial Toxins; D002942:Circular Dichroism; D003001:Cloning, Molecular; D016207:Cytokines / Q000378:metabolism; D004768:Enterotoxins / Q000235:genetics* / Q000276:immunology / Q000502:physiology*; D005260:Female; D005998:Glycine / Q000235:genetics; D000949:Histocompatibility Antigens Class II / Q000235:genetics* / Q000502:physiology; D006801:Humans; D019811:Hydroxylamine; D006898:Hydroxylamines / Q000494:pharmacology; D015151:Immunoblotting; D007963:Leukocytes, Mononuclear; D051379:Mice; D008807:Mice, Inbred BALB C; D008958:Models, Molecular; D016296:Mutagenesis; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D017354:Point Mutation; D017422:Sequence Analysis, DNA; D018089:Superantigens / Q000235:genetics* / Q000502:physiology
D000907:Antibodies, Bacterial; D001427:Bacterial Toxins; D016207:Cytokines; D004768:Enterotoxins; D000949:Histocompatibility Antigens Class II; D006898:Hydroxylamines; D018089:Superantigens; C030107:enterotoxin F, Staphylococcal; D019811:Hydroxylamine; D001224:Aspartic Acid; D005998:Glycine