Modulation of structured polypeptide specificity
The invention describes a method for selecting a polypeptide ligand having a desired level of specificity for a target, wherein the polypeptide ligand comprises a polypeptide comprising at least three reactive groups, separated by at least two loop sequences, and a molecular scaffold which forms covalent bonds with the reactive groups of the polypeptide such that at least two polypeptide loops are formed on the molecular scaffold, comprising (i) screening at least two different sets of polypeptide ligands against the target, and selecting one or more ligands from each library which interact with the target; (ii) comparing the activity of the selected ligands with one or more paralogues or orthologues of the target; and (iii) further selecting one or more ligands according to their activity towards said one or more paralogues or orthologues; wherein said two or more different sets of ligands differ in the length of the polypeptide loops formed on the molecular scaffold.
1. A polypeptide ligand comprising a polypeptide comprising at least three reactive groups, separated by at least two loop sequences and comprising the sequence AC(X) 4 JC(X) 5 CG [SEQ ID No: 1], wherein X is a random natural amino acid, A is Alanine, C is Cysteine, G is Glycine, and J is N-methyl Arginine, Homo-Arginine, or Guanidylphenylalanine; and a molecular scaffold which forms covalent bonds with the reactive groups of the polypeptide such that at least two polypeptide loops are formed on the molecular scaffold and the polypeptide ligand is resistant to protease degradation.
2. The polypeptide ligand according to claim 1 , wherein the polypeptide ligand comprises only two loops.
3. The polypeptide ligand according to claim 1 , wherein the polypeptide ligand comprises three loops.
4. The polypeptide ligand according to claim 1 , which is specific for a protease.
5. The polypeptide ligand according to claim 4 , wherein the protease is plasma kallikrein.