1. Mutation of surface residues to promote crystallization of activated factor XI as a complex with benzamidine: an essential step for the iterative structure-based design of factor XI inhibitors.
- Author
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Jin L, Pandey P, Babine RE, Weaver DT, Abdel-Meguid SS, and Strickler JE
- Subjects
- Binding Sites, Blood Coagulation Factors chemistry, Catalysis, Catalytic Domain, Crystallography, X-Ray, DNA, Complementary metabolism, Humans, Hydrogen Bonding, Inhibitory Concentration 50, Ligands, Macromolecular Substances chemistry, Models, Molecular, Mutagenesis, Mutagenesis, Site-Directed, Peptides chemistry, Pichia metabolism, Protein Binding, Protein Conformation, Recombinant Proteins chemistry, Serine chemistry, Benzamidines chemistry, Factor XI antagonists & inhibitors, Factor XIa chemistry, Factor XIa genetics, Mutation
- Abstract
Activated factor XI (FXIa) is a key enzyme in the amplification phase of the blood-coagulation cascade. Thus, a selective FXIa inhibitor may have lesser bleeding liabilities and provide a safe alternative for antithrombosis therapy to available drugs on the market. In a previous report, the crystal structures of the catalytic domain of FXIa (rhFXI(370-607)) in complex with various ecotin mutants have been described. However, ecotin forms a matrix-like interaction with rhFXI(370-607) and is impossible to displace with small-molecule inhibitors; ecotin crystals are therefore not suitable for iterative structure-based ligand design. In addition, rhFXI(370-607) did not crystallize in the presence of small-molecule ligands. In order to obtain the crystal structure of rhFXI(370-607) with a weak small-molecule ligand, namely benzamidine, several rounds of surface-residue mutation were implemented to promote crystal formation of rhFXI(370-607). A quadruple mutant of rhFXI(370-607) (rhFXI(370-607)-S434A,T475A,C482S,K437A) readily crystallized in the presence of benzamidine. The benzamidine in the preformed crystals was easily exchanged with other FXIa small-molecule inhibitors. These crystals have facilitated the structure-based design of small-molecule FXIa inhibitors.
- Published
- 2005
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