ProteaseSpots™
JPT Peptide Technologies offers a new and efficient way to study protease activities and substrate specificities. ProteaseSpots™ are custom synthesized protease substrates that are immobilized on cellulose discs. The discs are delivered ready-to use in microtiter plates and are incubated with your protease. Subsequent fluorescent detection can be performed after several time points.
Benefits of the ProteaseSpots™ System
- Screen large numbers of substrates economically
- You specify the individual substrate sequences
- Obtain semi-kinetic data
- Easy handling in 96-well plates
Applications for ProteaseSpots™
- Detection of protein cleavage sites utilizing peptide scans
- Identification of novel substrates with de novo libraries
- Measurement of protease activities with known substrates
- Characterization of cleavage sites through substitutional analyses
Figure: The protease assay is based on peptides bound to small cellulose discs in 96-well microtiter plates.
These peptides bear a fluorescent moiety at the N-terminus. After incubation with the active protease, aliquots are transferred to an additional plate for detection of the N-terminally labeled peptide fragment (excitation: 325 nm, emission: 420 nm).
Figure: Fluorescence intensities of a caspase-3 substrate VDQMDGW single site L-amino acid substitutional analysis.
This method identifies new or improved substrates.
Each residue in the sequence (rows) was replaced by all 20 amino acids (columns).
Figure: Fluorescence intensity of trypsin cleaved peptides derived from hen egg lysozyme.
The entire sequence was synthesized using 7-mer overlapping peptides shifted by three amino acids.
Related products and services
Protease Substrate Sets
Internally quenched (Dabcyl/EDANS) pre-made protease substrate collections ready-to-screen in microtiter plates.
Protease Peptide Microarrays
Compilation of protease substrate peptides on ready-to-use PepStar™ peptide microarrays.
Protease Substrate Identification
Profiling service for protease using our high-content PepStar™ peptide microarrays.
Selected References:
- Identification of Candidate Substrates for Ectodomain Shedding by the Metalloprotease-Disintegrin ADAM8
Naus et al., Biol Chem. 2006 (2006) (abstract) - Screening a Combinatorial Peptide Library to Develop a Human Glandular Kallikrein-2 Activated Prodrug as Targeted Therapy for Prostate Cancer
Janssen et al, Mol. Cancer Ther. (2004) (abstract) - Processing of the Human Transferrin Receptor at Distinct Positions within the Stalk Region by Neutrophil Elastase and Cathepsin
Kaup et al., Biol. Chem. (2002) (abstract) - Escherichia Coli DegP Protease Cleaves Between Paired Hydrophobic Residues in a Natural Substrate: the PapA Pilin
Jones et al, J. Bacteriol. (2002) (abstract)
More references under JPT Publications/Literature
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