Evidence for Novel Action at the Cell-Binding Site of Human Angiogenin Revealed by Heteronuclear NMR Spectroscopy, insilico and invivo Studies

A member of the ribonucleaseA superfamily, human angiogenin (hAng) is a potent angiogenic factor. Heteronuclear NMR spectroscopy combined with induced-fit docking revealed a dual binding mode for the most antiangiogenic compound of a series of ribofuranosyl pyrimidine nucleosides that strongly inhibit hAng's angiogenic activity invivo. While modeling suggests the potential for simultaneous binding of the inhibitors at the active and cell-binding sites, NMR studies indicate greater affinity for the cell-binding site than for the active site. Additionally, molecular dynamics simulations at 100ns confirmed the stability of binding at the cell-binding site with the predicted protein-ligand interactions, in excellent agreement with the NMR data. This is the first time that a nucleoside inhibitor is reported to completely inhibit the angiogenic activity of hAng invivo by exerting dual inhibitory activity on hAng, blocking both the entrance of hAng into the cell and its ribonucleolytic activity.