The rapid endopeptidase profiling library (REPLi) is a generic screening tool that overcomes known hurdles to identifying the cleavage site requirements for protease substrates, which are viewed as possible drug targets in disease treatment.
The peptide library (PepSet) consists of fluorescently labelled tripeptide analogs spanning the sequence surrounding the cleavage site of the substrate. With just 512 distinct pools containing eight peptides in each, the library can be relatively small while still representing the residue requirements for the largest number of proteases.
The variable central core of each peptide is flanked with multiple Gly residues and an additional two Lys residues are added at the C-terminus, to confer adequate solubility to peptides bearing hydrophobic variable sequences. Potentially problematic amino acids have been removed from the selection set, while certain others are grouped in matching pairs (Ala + Val, Arg + Lys, Asp + Glu, Asn + Gln, Leu + lle, Ser + Thr, Phe + Tyr). This choice of amino acid partners ensures that maximum SAR information can be derived from an initial result and greatly simplifies the deconvolution steps.
The complete peptide library is synthesised using Mimotopes’ PepSets SynPhase Lantern technology and is provided for direct usage in 96-well plate format.
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