title: Characterization of the Staphylococcal enterotoxin A: Vβ receptor interaction using human receptor fragments engineered for high affinity.
abstract: Staphylococcal food poisoning is a gastrointestinal disorder caused by the consumption of food containing Staphylococcal enterotoxins. Staphylococcal enterotoxin A (SEA) is the most common enterotoxin recovered from food poisoning outbreaks in the USA. In addition to its enteric activity, SEA also acts as a potent superantigen through stimulation of T cells, although less is known about its interactions than the superantigens SEB, SEC and toxic shock syndrome toxin-1. To understand more about SEA:receptor interactions, and to develop toxin-detection systems for use in food testing, we engineered various SEA-binding receptor mutants. The extracellular domain of the receptor, a variable region of the beta chain (Vβ22) of the T-cell receptor, was engineered for stability as a soluble protein and for high affinity, using yeast-display technology. The highest affinity mutant was shown to bind SEA with a Kd value of 4 nM. This was a 25 000-fold improvement in affinity compared with the wild-type receptor, which bound to SEA with low affinity (Kd value of 100 µM), similar to other superantigen:Vβ interactions. The SEA:Vβ interface was centered around residues within the complementarity determining region 2 loop. The engineered receptor was specific for SEA, in that it did not bind to two other closely related enterotoxins SEE or SED, providing information on the SEA residues possibly involved in the interaction. The specificity and affinity of these high-affinity Vβ proteins also provide useful agents for the design of more sensitive and specific systems for SEA detection.
authors.affiliation: Department of Biochemistry, University of Illinois, Urbana, IL 61801, USA.
authors.forename: P
authors.initials: P
authors.lastname: Sharma
authors.forename: S
authors.initials: S
authors.lastname: Postel
authors.forename: E J
authors.initials: EJ
authors.lastname: Sundberg
authors.forename: D M
authors.initials: DM
authors.lastname: Kranz
mesh_terms: D000595:Amino Acid Sequence; D061505:Cell Surface Display Techniques; D004768:Enterotoxins / Q000032:analysis / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding / Q000235:genetics; D055550:Protein Stability; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012680:Sensitivity and Specificity; D016415:Sequence Alignment
chemical_list: D004768:Enterotoxins; D016693:Receptors, Antigen, T-Cell, alpha-beta; D011994:Recombinant Proteins; C026066:enterotoxin A, Staphylococcal
keywords: Staphylococcal enterotoxin A (SEA); directed evolution; food poisoning; yeast display