The presence of multiple protein binding sites on single heparin chains could not allow them to bind non-covalently to oligopeptide components of tissue scaffolds while retaining growth factor affinity.  However, the extent of this oligopeptide coupling interferes with conjugate protein binding is unknown.  To elucidate such simultaneous specific and non-specific interactions, we examined a well-defined ternary system comprising fibroblast growth factor (FGF), tetralysine (K4), with a heparin decamer (dp10) acting as a non-covalent coupler.  Electrospray ionization mass spectrometry was used to assess binding affinities and complex stoichiometries as a function of ionic strength I for dp10-K4 and FGF-dp10.  The electrostatic nature of the former was evident from the decrease in binding above I = 40 mM, while the stronger binding for FGF-dp10 can be inferred from its greater salt resistance.  Remarkably, dp10 can bind two FGF, but only one tetralysine.  The limited binding of K4 to dp10 suggests that the tetralysine might not block growth factor binding; the 1:1:1 ternary complex is indeed observed.  The salt resistance of the FGF-heparin and K4-heparin complexes shows that the former interaction is stronger.  Observation of the compositional distribution of free and FGF-bound dp10 suggest a correlation between heparin degree of sulfonation and FGF affinity, indicating a more nuanced approach than the use of heterogeneous native heparin for growth factor sequestration and release.

Bridging aFGF to a cationic oligopeptide via specific and nonspecific interactions with a heparin decamer

246th National Meeting (2013)

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