The NMR solution structure of recombinant RGD-hirudin

The solution structure of a new recombinant RGD-hirudin, which has the activities of anti-thrombin and anti-platelet aggregation, was determined by 1H nuclear magnetic resonance spectroscopy and compared with the conformations of recombinant wild-type hirudin and hirudin (variant 2, Lys47) of the hirudin thrombin complex. On the basis of total 1284 distance and dihedral angle constraints derived from a series of NMR spectra, 20 conformers were computed with ARIA/CNS programs. The structure of residues 3–30 and 37–48 form a molecular core with two antiparallel β-sheets as the other two hirudins. However, significant differences were found in the surface electrostatic charge distributions among the three hirudins, especially in the RGD segment of recombinant RGD-hirudin. This difference may be greatly beneficial to its additional function of anti-platelet aggregation. The difference in extended C-terminal makes its both ionic and hydrophobic interactions with the fibrinogen recognition exosite of thrombin more effective.