A mutation at histidine residue 135 of toxic shock syndrome toxin yields an immunogenic protein with minimal toxicity.
Structure-function studies have revealed that the region between amino acids 115 and 141 of toxic shock syndrome toxin 1 (TSST-1) constitutes a biologically active domain. A critical residue appears to be histidine 135, since a site-directed mutation that alters the histidine to alanine (H135A) results in a loss of mitogenic activity and an absence of toxicity as measured in a rabbit infection model of toxic shock syndrome. We have characterized the mutant toxin further and report here on its immunogenic activity in rabbits and on the protective ability of mutant-specific antibodies in two animal models of toxin-mediated shock. Antibodies raised in rabbits by immunization with the purified H135A are fully cross-reactive with staphylococcal TSST-1 and wild-type recombinant TSST-1 (rTSST-1) expressed in Escherichia coli. The H135A antibodies neutralized the mitogenic activity for murine splenic T cells equally well as did TSST-1-specific polyclonal and monoclonal antibodies. In addition, the H135A antibodies blocked the production of tumor necrosis factor by spleen cells stimulated with rTSST-1. The toxicities of rTSST-1 and H135A were compared in D-galactosamine (D-GalNH2)-sensitized MRL-lpr/lpr mice. The nontoxicity of H135A was confirmed in this murine model of superantigen-induced septic shock. No toxicity of H135A was demonstrable at doses of 60 micrograms, while doses of rTSST-1 as low as 2 micrograms caused significant mortality within 24 to 72 h after challenge. Furthermore, subsequent to challenge of mice with H135A, no elevation in the serum levels of interleukin-2 or tumor necrosis factor was measurable. Passive immunization with H135A antibodies also protected MRL-lpr/lpr mice against lethal challenge with rTSST-1. Finally, rabbits actively immunized with purified H135A did not succumb to infection with a transformed strain of Staphylococcus aureus expressing rTSST-1. Additional animal studies will be required to confirm the immunizing potential of H135A and the efficacy of H135A antibodies as a neutralizing antitoxin.
Bonventre
P F
PF
Department of Molecular Genetics, Biochemistry, and Microbiology, University of Cincinnati College of Medicine, Ohio 45267.
Heeg
H
H
Edwards
C K
CK
Cullen
C M
CM
D000818:Animals; D000907:Antibodies, Bacterial / Q000276:immunology; D001427:Bacterial Toxins; D003429:Cross Reactions; D004768:Enterotoxins / Q000737:chemistry / Q000276:immunology* / Q000633:toxicity; D005260:Female; D006639:Histidine; D008213:Lymphocyte Activation / Q000187:drug effects; D008297:Male; D051379:Mice; D008817:Mice, Mutant Strains; D008934:Mitogens; D016297:Mutagenesis, Site-Directed; D011817:Rabbits; D012772:Shock, Septic / Q000517:prevention & control*; D013329:Structure-Activity Relationship; D018089:Superantigens; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis
D000907:Antibodies, Bacterial; D001427:Bacterial Toxins; D004768:Enterotoxins; D008934:Mitogens; D018089:Superantigens; D014409:Tumor Necrosis Factor-alpha; C030107:enterotoxin F, Staphylococcal; D006639:Histidine