Blood Journal
Leading the way in experimental and clinical research in hematology

Building better fibrin knob mimics: an investigation of synthetic fibrin knob peptide structures in solution and their dynamic binding with fibrinogen/fibrin holes

  1. Sarah E. Stabenfeldt1,
  2. J. Jared Gossett2, and
  3. Thomas H. Barker1,*
  1. 1 W.H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology / Emory University, Atlanta, GA, United States;
  2. 2 School of Biology, School of Computational Science and Engineering, Georgia Institute of Technology, Atlanta, GA, United States
  1. * Corresponding author; email: thomas.barker{at}bme.gatech.edu

Abstract

Fibrin polymerizes via non-covalent and dynamic association of thrombin-exposed 'knobs' with complementary 'holes'. Synthetic knob peptides have received significant interest as a means for understanding fibrin assembly mechanisms and inhibiting fibrin polymerization. Nevertheless, the inability to crystallize short peptides significantly limits our understanding of knob peptide structural features that regulate dynamic knob:hole interactions. In this study, we utilized molecular simulations to generate the first predicted structure(s) of synthetic knobs in solution prior to fibrin hole engagement. Combining Surface Plasmon Resonance (SPR), we explored the role of structural and electrostatic properties of knob 'A' mimics in regulating knob:hole binding kinetics. SPR results illustrated that association rates were most profoundly affected by the presence of both additional prolines as well as charged residues in the 6-7th positions. Importantly, analyzing the structural dynamics of the peptides through simulation indicated that the 3Arg side chain orientation and peptide backbone stability each contribute significantly to functional binding. These findings provide insights into early fibrin protofibril assembly dynamics as well as establish essential design parameters for high affinity knob mimics that more efficiently compete for hole occupancy, parameters realized here through a novel knob mimic displaying a 10-fold higher association rate than current mimics.

  • Submitted November 3, 2009.
  • Accepted May 9, 2010.