title: Mutational effects on protein folding stability and antigenicity: the case of streptococcal pyrogenic exotoxin A.
abstract: The influence of mutationally induced changes in protein folding on development of effective neutralizing antibodies during vaccination remains largely unexplored. In this study, we probed how mutational substitutions of streptococcal pyrogenic exotoxin A (SPEA), a model bacterial superantigen, affect native conformational stability and antigenicity. Stability changes for the toxin variants were determined using circular dichroism and fluorescence measurements, and scanning calorimetry. Self-association was assayed by dynamic light scattering. Inactivated SPEA proteins containing particular combinations of mutations elicited antibodies in HLA-DQ8 transgenic mice that neutralized SPEA superantigenicity in vitro, and protected animals from lethal toxin challenge. However, a highly destabilized cysteine-free mutant of SPEA did not provide effective immunity, nor did an irreversibly denatured version of an otherwise effective mutant protein. These results suggest that protein conformation plays a significant role in generating effective neutralizing antibodies to this toxin, and may be an important factor to consider in vaccine design.
authors.affiliation: United States Army Medical Research Institute of Infectious Diseases, Department of Cell Biology and Biochemistry, 1425 Porter Street, Frederick, MD 21702, USA. john.carra@det.amedd.army.mil
authors.forename: John H
authors.initials: JH
authors.lastname: Carra
authors.forename: Brent C
authors.initials: BC
authors.lastname: Welcher
authors.forename: Rowena D
authors.initials: RD
authors.lastname: Schokman
authors.forename: Chella S
authors.initials: CS
authors.lastname: David
authors.forename: Sina
authors.initials: S
authors.lastname: Bavari
mesh_terms: D000818:Animals; D001426:Bacterial Proteins; D019281:Dimerization; D005098:Exotoxins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D006358:Hot Temperature; D008565:Membrane Proteins; D051379:Mice; D008822:Mice, Transgenic; D017510:Protein Folding; D013329:Structure-Activity Relationship; D013816:Thermodynamics; D014612:Vaccines / Q000276:immunology*
chemical_list: D001426:Bacterial Proteins; D005098:Exotoxins; D008565:Membrane Proteins; C117980:SpeA protein, Streptococcus pyogenes; D014612:Vaccines; C027951:erythrogenic toxin