The interaction between heparin and Lys49 phospholipase A2 reveals the natural binding of heparin on the enzyme.

We have studied at a molecular level the interaction of heparins on bothropstoxin-I (BthTx-I), a phospholipase A2 toxin. The protein was monitored using gel filtration chromatography, dynamic light scattering (DLS), circular dichroism (CD), attenuated total reflectance Fourier transform infrared (ATR-FTIR) and intrinsic tryptophan fluorescence emission (ITFE) spectroscopy. The elution profile of the protein presents a displacement of the protein peak to larger complexes when interacting with higher concentration of heparin. The DLS results shows two Rh at a molar ratio of 1, one to the distribution of the protein and the second for the action of heparin on BthTx-I structures, and a large distribution with the increase of protein. The interaction is accompanied by significant changes in the CD spectra, showing two common features: a decrease in signal at 208 nm (3 and 6 kDa heparins) and an isodichroic point near 226 nm (3 kDa heparin). FTIR spectra indicate that only a few amino acid residues are involved in this interaction. Alterations in the ITFE by binding heparins suggest that the initial binding occurs on the ventral face of BthTx-I. Together, these results add an experimental and structural basis on the action mechanism of the heparins over the phospholipases A2 and provide a molecular model to elucidate the interaction of the enzyme-heparin complex at a molecular level.