Rational design and selection of bivalent peptide ligands of thrombin incorporating P4-P1 tetrapeptide sequences: from good substrates to potent inhibitors

Protein Engineering, Design & Selection : PEDS
Zhengding SuFeng Ni

Abstract

The tetrapeptide Phe-Asn-Pro-Arg is a structurally optimized sequence for binding to the active site of thrombin. By conjugating this tetrapeptide or some variants to a C-terminal fragment of hirudin, we were able to generate a series of new bivalent inhibitors of thrombin containing only genetically encodable natural amino acids. We found that synergistic binding to both the active site and an exosite of thrombin can be enhanced through substitutions of amino acid residues at the P3 and P3' sites of the active-site directed sequence, Phe(P4)-Xaa(P3)-Pro(P2)-Arg(P1)-Pro(P1')-Gln(P2')-Yaa(P3'). Complementary to rational design, a phage library was constructed to explore further the residue requirements at the P4, P3 and P3' sites for bivalent and optimized two-site binding. Very significantly, panning of the phage library has led to thrombin-inhibitory peptides possessing strong anti-clotting activities in the low nanomolar range and yet interfering only partially the catalytic active site of thrombin. Modes of action of the newly discovered bivalent inhibitors are rationalized in light of the allosteric properties of thrombin, especially the interplay between the proteolytic action and regulatory binding occurring at thrombin s...Continue Reading

Citations

Mar 10, 2010·Proceedings of the National Academy of Sciences of the United States of America·Laurent VolponKatherine L B Borden
Nov 29, 2011·International Journal of Cancer. Journal International Du Cancer·Rana FilfilAnne E G Lenferink
Oct 20, 2006·Protein Expression and Purification·Hongjian LiJames Z Su
Jun 12, 2010·Angewandte Chemie·Hongjian LiZhengding Su
Aug 13, 2020·Circulation Research·Dong Heon LeeAndrew D Johnson
Mar 25, 2010·Langmuir : the ACS Journal of Surfaces and Colloids·Jocelyne AdjémianChristophe Demaille

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