Characterization of T Cell Receptors Engineered for High Affinity Against Toxic Shock Syndrome Toxin-1

Buonpane, Rebecca A.; Moza, Beenu; Sundberg, Eric J.; Kranz, David M.

doi:10.1016/j.jmb.2005.08.041

Cited by 49 publications

(84 citation statements)

References 35 publications

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“…To further explore the molecular basis of the high-affinity binding, we generated the six reversion mutants of the 4-5 protein and examined them in the yeast display system. This approach allows more rapid analysis than expression of soluble mutants, because titrations of yeast-displayed mutants can be performed without the need for protein purification (10). Flow cytometry-based titrations of each mutant with SpeA (Fig.…”

Section: Resultsmentioning

confidence: 99%

A Single, Engineered Protein Therapeutic Agent Neutralizes Exotoxins from BothStaphylococcus aureusandStreptococcus pyogenes

Wang

Mattis

Sundberg

et al. 2010

Clin Vaccine Immunol

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Staphylococcus aureus and Streptococcus pyogenes secrete exotoxins that act as superantigens, proteins that cause hyperimmune reactions by binding the variable domain of the T-cell receptor beta chain (V␤), leading to stimulation of a large fraction of the T-cell repertoire. To develop potential neutralizing agents, we engineered V␤ mutants with high affinity for the superantigens staphylococcal enterotoxin B (SEB), SEC3, and streptococcal pyrogenic exotoxin A (SpeA). Unexpectedly, the high-affinity V␤ mutants generated against SEB cross-reacted with SpeA to a greater extent than they did with SEC3, despite greater sequence similarity between SEB and SEC3. Likewise, the V␤ mutants generated against SpeA cross-reacted with SEB to a greater extent than with SEC3. The structural basis of the high affinity and cross-reactivity was examined by single-site mutational analyses. The cross-reactivity seems to involve only one or two toxin residues. Soluble forms of the cross-reactive V␤ regions neutralized both SEB and SpeA in vivo, suggesting structure-based strategies for generating high-affinity neutralizing agents that can cross-react with multiple exotoxins.

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Section: Resultsmentioning

confidence: 99%

A Single, Engineered Protein Therapeutic Agent Neutralizes Exotoxins from BothStaphylococcus aureusandStreptococcus pyogenes

Wang

Mattis

Sundberg

et al. 2010

Clin Vaccine Immunol

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“…Binding to the TCR Vβ CDR2 loop is a requirement for all bacterial SAGs, and the proportion of the SAG-TCR interface that is contributed by the CDR2 loop is invariably the greatest in any SAG-TCR complex, relative to any other single hypervariable or framework region. Involvement of Vβ domain regions beyond the CDR2 loop, however, plays a significant role in the TCR Vβ domain specificity and cross-reactivity of a SAG [38,49,54]. SEK and TSST-1 engage one or more framework region apical loops, at the expense of contacting the hypervariable elements.…”

Section: Sag-tcr Specificity and Cross-reactivitymentioning

confidence: 99%

“…Together, these affinities allow the overall MHC/SpeC/ TCR complex to achieve an affinity within the range for efficient T cell activation. The affinities of TSST-1 for pMHC and TCR are 1 μM [37] and 0.6 μM [54], respectively. The overall sub-μM affinity of the MHC/TSST-1/TCR complex is thus within the range exhibited by most pMHC/TCR interactions [57].…”

Section: Superantigen-mediated T Cell Signaling Complexesmentioning

confidence: 99%

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TCR recognition of peptide/MHC class II complexes and superantigens

Sundberg

Deng

Mariuzza

2007

Seminars in Immunology

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SummaryMajor histocompatibility complex (MHC) class II molecules display peptides to the T cell receptor (TCR). The ability of the TCR to discriminate foreign from self peptides presented by MHC molecules is a requirement of an effective adaptive immune response. Dysregulation of this molecular recognition event often leads to a disease state. Recently, a number of structural studies have provided significant insight into several such dysregulated interactions between peptide/MHC complexes and TCR molecules. These include TCR recognition of self peptides, which results in autoimmune reactions, and of mutant self-peptides, common in the immunosurveillance of tumors, as well as the engagement of TCRs by superantigens, a family of bacterial toxins responsible for toxic shock syndrome.

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“…In this work, we explored the use of engineered soluble receptor proteins that were developed as a way to neutralize SEs during the disease process (9)(10)(11)(12). The SE binds to the T-cell receptor (TCR) variable region of the ␤ chain (V␤) (TCR-V␤) on a T cell and to the class II major histocompatibility complex (MHC) molecule on an antigen-presenting cell.…”

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Novel Platform for the Detection of Staphylococcus aureus Enterotoxin B in Foods

Tallent

DeGrasse

Wang

et al. 2013

Appl Environ Microbiol

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bStaphylococcal contamination of food products and staphylococcal food-borne illnesses continue to be a problem worldwide. Screening of food for the presence of Staphylococcus aureus and/or enterotoxins using traditional methods is laborious. Reliable and rapid multiplex detection methods from a single food extract or culture supernatant would simplify testing. A fluorescence-based cytometric bead array was developed for the detection of staphylococcal enterotoxin B (SEB), using magnetic microspheres coupled with either an engineered, enterotoxin-specific V␤ domain of the T-cell receptor (V␤-TCR) or polyclonal antibodies. The binding affinity of the V␤-TCR for SEB has been shown to be in the picomolar range, comparable to the best monoclonal antibodies. The coupled beads were validated with purified enterotoxins and tested in a variety of food matrices spiked with enterotoxins. The V␤-TCR or antibody was shown to specifically bind SEB in four different food matrices, including milk, mashed potatoes, vanilla pudding, and cooked chicken. The use of traditional polyclonal antibodies and V␤-TCR provides a redundant system that ensures accurate identification of the enterotoxin, and the use of labeled microspheres permits simultaneous testing of multiple enterotoxins from a single sample. Staphylococcus aureus is among the top five common pathogens associated with food-borne illness nationwide. The Centers for Disease Control and Prevention (CDC) estimates that there are about 240,000 illnesses with 1,000 hospitalizations and 6 deaths associated with staphylococcal food poisoning (SFP) annually (1). SFP is directly linked to small (25-to 30-kDa) exotoxins, known as staphylococcal enterotoxins (SEs), that are heat resistant, tolerate low pH, and often persist long after the microbe has been rendered nonviable. SEs are superantigens that induce nonspecific T-cell activation, which can rapidly cascade to a massive release of inflammatory mediators and may lead to toxic shock (2, 3). SFP occurs when improperly handled food contaminated with as little as 100 ng of SE is consumed. SFP is marked by severe gastrointestinal symptoms such as emesis, diarrhea, and/or abdominal pain after a 4-h incubation period (4).Suspect food products are typically analyzed by using traditional culture techniques followed by identification of enterotoxigenic staphylococcal strains. Food extracts positive for bacteria may then be evaluated for the presence of preformed SEs (5). A recent outbreak in Japan from milk contaminated with staphylococcal enterotoxin A (SEA) illustrated the challenges of conventional methods, since viable bacteria were not detected (6, 7). PCR methods would be a useful screening tool for samples that are culture negative and enterotoxin positive (8). PCR methods are sensitive and can detect low levels of gene targets; however, this does not verify the presence of expressed proteins, and it is unacceptable as a definitive method for SE detection in the regulatory arena. Therefore, immunological techniques using either m...

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Characterization of T Cell Receptors Engineered for High Affinity Against Toxic Shock Syndrome Toxin-1

Cited by 49 publications

References 35 publications

A Single, Engineered Protein Therapeutic Agent Neutralizes Exotoxins from BothStaphylococcus aureusandStreptococcus pyogenes

A Single, Engineered Protein Therapeutic Agent Neutralizes Exotoxins from BothStaphylococcus aureusandStreptococcus pyogenes

TCR recognition of peptide/MHC class II complexes and superantigens

Novel Platform for the Detection of Staphylococcus aureus Enterotoxin B in Foods

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