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A single, engineered protein therapeutic agent neutralizes exotoxins from both Staphylococcus aureus and Streptococcus pyogenes.

ABSTRACT: 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.

SUBMITTER: Wang N

PROVIDER: S-EPMC2976085 | biostudies-literature | 2010 Nov

REPOSITORIES: biostudies-literature

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A single, engineered protein therapeutic agent neutralizes exotoxins from both Staphylococcus aureus and Streptococcus pyogenes.

Wang Ningyan N Mattis Daiva M DM Sundberg Eric J EJ Schlievert Patrick M PM Kranz David M DM

Clinical and vaccine immunology : CVI 20100922 11

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 (Vβ), leading to stimulation of a large fraction of the T-cell repertoire. To develop potential neutralizing agents, we engineered Vβ mutants with high affinity for the superantigens staphylococcal enterotoxin B (SEB), SEC3, and streptococcal pyrogenic exotoxin A (SpeA). Unexpectedly, the high-affi ...[more]

PMID: 20861327

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Project description:Identification of genetic polymorphisms causing increased antibiotic resistance in bacterial pathogens traditionally has proceeded from observed phenotype to defined mutant genotype. The availability of large collections of microbial genome sequences that lack antibiotic susceptibility metadata provides an important resource and opportunity to obtain new information about increased antimicrobial resistance by a reverse genotype-to-phenotype bioinformatic and experimental workflow. We analyzed 26,465 genome sequences of Streptococcus pyogenes, a human pathogen causing 700 million infections annually. The population genomic data identified amino acid changes in penicillin-binding proteins 1A, 1B, 2A, and 2X with signatures of evolution under positive selection as potential candidates for causing decreased susceptibility to β-lactam antibiotics. Construction and analysis of isogenic mutant strains containing individual amino acid replacements in penicillin-binding protein 2X (PBP2X) confirmed that the identified residues produced decreased susceptibility to penicillin. We also discovered the first chimeric PBP2X in S. pyogenes and show that strains containing it have significantly decreased β-lactam susceptibility. The novel integrative reverse genotype-to-phenotype strategy presented is broadly applicable to other pathogens and likely will lead to new knowledge about antimicrobial agent resistance, a massive public health problem worldwide. IMPORTANCE The recent demonstration that naturally occurring amino acid substitutions in Streptococcus pyogenes PBP2X are sufficient to cause severalfold reduced susceptibility to multiple β-lactam antibiotics in vitro raises the concern that these therapeutic agents may become compromised. Substitutions in PBP2X are common first-step mutations that, with the incremental accumulation of additional adaptive mutations within the PBPs, can result in high-level resistance. Because β-lactam susceptibility testing is not routinely performed, the nature and extent of such substitutions within the PBPs of S. pyogenes are poorly characterized. To address this knowledge deficit, polymorphisms in the PBPs were identified among the most comprehensive cohort of S. pyogenes genome sequences investigated to date. The mutational processes and selective forces acting on the PBPs were assessed to identify specific substitutions likely to influence β-lactam susceptibility and to evaluate factors posited to be impediments to resistance emergence.

Dataset Information

A single, engineered protein therapeutic agent neutralizes exotoxins from both Staphylococcus aureus and Streptococcus pyogenes.

Publications

A single, engineered protein therapeutic agent neutralizes exotoxins from both Staphylococcus aureus and Streptococcus pyogenes.

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