A peptide-based fluorescence resonance energy transfer assay for Bacillus anthracis lethal factor protease
AUTOR(ES)
Cummings, Richard T.
FONTE
The National Academy of Sciences
RESUMO
A fluorescence resonance energy transfer assay has been developed for monitoring Bacillus anthracis lethal factor (LF) protease activity. A fluorogenic 16-mer peptide based on the known LF protease substrate MEK1 was synthesized and found to be cleaved by the enzyme at the anticipated site. Extension of this work to a fluorogenic 19-mer peptide, derived, in part, from a consensus sequence of known LF protease targets, produced a much better substrate, cleaving approximately 100 times more efficiently. This peptide sequence was modified further on resin to incorporate donor/quencher pairs to generate substrates for use in fluorescence resonance energy transfer-based appearance assays. All peptides cleaved at similar rates with signal/background ranging from 9–16 at 100% turnover. One of these substrates, denoted (Cou)Consensus(K(QSY-35)GG)-NH2, was selected for additional assay optimization. A plate-based assay requiring only low nanomolar levels of enzyme was developed for screening and inhibitor characterization.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=124449Documentos Relacionados
- Synthetic peptide-based enzyme-linked immunosorbent assay for serodiagnosis of visceral leishmaniasis.
- Peptide-Based OspC Enzyme-Linked Immunosorbent Assay for Serodiagnosis of Lyme Borreliosis
- De novo designed peptide-based amyloid fibrils
- Cell-free tumor antigen peptide-based cancer vaccines
- Loligomers: design of de novo peptide-based intracellular vehicles.
