RESUMO
The two subunits of core binding factor (Runx1 and CBFbeta) play critical roles in hematopoiesis and are frequent targets of chromosomal translocations found in leukemia. The binding of the CBFbeta-smooth muscle myosin heavy chain (SMMHC) fusion protein to Runx1 is essential for leukemogenesis, making this a viable target for treatment. We have developed inhibitors with low micromolar affinity which effectively block binding of Runx1 to CBFbeta. NMR-based docking shows that these compounds bind to CBFbeta at a site displaced from the binding interface for Runx1, that is, these compounds function as allosteric inhibitors of this protein-protein interaction, a potentially generalizable approach. Treatment of the human leukemia cell line ME-1 with these compounds shows decreased proliferation, indicating these are good candidates for further development.
Assuntos
Sítio Alostérico , Proliferação de Células/efeitos dos fármacos , Subunidade alfa 2 de Fator de Ligação ao Core/antagonistas & inibidores , Subunidade beta de Fator de Ligação ao Core/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Leucemia/patologia , Sítios de Ligação , Linhagem Celular Tumoral , Subunidade alfa 2 de Fator de Ligação ao Core/química , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Subunidade beta de Fator de Ligação ao Core/química , Subunidade beta de Fator de Ligação ao Core/metabolismo , Inibidores Enzimáticos/química , Transferência Ressonante de Energia de Fluorescência , Hematopoese/genética , Hematopoese/fisiologia , Humanos , Leucemia/metabolismo , Espectroscopia de Ressonância Magnética , Cadeias Pesadas de Miosina/química , Cadeias Pesadas de Miosina/metabolismo , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/metabolismo , Miosinas de Músculo Liso/química , Miosinas de Músculo Liso/metabolismo , Translocação Genética/genética , Translocação Genética/fisiologiaRESUMO
2,2,2-Trifluoroethanol (TFE) is widely used to induce helix formation in peptides and proteins, but the mechanism behind this effect is still poorly understood. Several recent papers have proposed that TFE acts by selectively desolvating the peptide backbone groups of the helix state. Infrared (IR) spectroscopy of the amide I band of polypeptides can be used to probe both secondary structure and backbone solvation, making this technique well suited for addressing the effect of TFE on polypeptide conformation. In this paper, we report the IR spectra as a function of TFE concentration for an alanine-rich peptide based on the repeat (AAKAA)(n)(). The IR spectra confirm that TFE desolvates the helical state of the peptide to a greater extent than the random coil state. Moreover, using a series of specifically (13)C-labeled peptides, the precise residues desolvated in the presence of TFE were identified. The residues most desolvated by TFE are the alanines located at position i - 4 in the sequence, where i is a lysine residue. This pattern of desolvation is consistent with molecular dynamics simulations which predict strong interactions between the lysine side chain at position n and the backbone carbonyl of the alanine at position i - 4. This is the first direct spectroscopic evidence of specific desolvation of helix backbone atoms in model alanine-rich peptides.
