Fibroblast activation protein (FAP) is a serine protease of undefined endopeptidase specificity implicated in tumorigenesis. To characterize FAP's P4-P′2 specificity, we synthesized intramolecularly quenched fluorescent substrate sets based on the FAP cleavage site in α2-antiplasmin (TSGP-NQ). FAP required substrates with Pro at P1 and Gly or d-amino acids at P2 and preferred small, uncharged amino acids at P3, but tolerated most amino acids at P 4, P′1 and P′2. These substrate preferences allowed design of peptidyl-chloromethyl ketones that inhibited FAP, but not the related protease, dipeptidyl peptidase-4. Thus, FAP is a narrow specificity endopeptidase and this can be exploited for inhibitor design. © 2006 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
Edosada, C. Y., Quan, C., Tran, T., Pham, V., Wiesmann, C., Fairbrother, W., & Wolf, B. B. (2006). Peptide substrate profiling defines fibroblast activation protein as an endopeptidase of strict Gly2-Pro1-cleaving specificity. FEBS Letters, 580(6), 1581–1586. https://doi.org/10.1016/j.febslet.2006.01.087