RESUMO
Knowledge of the structures formed by proteins and small ligands is of fundamental importance for understanding molecular principles of chemotherapy and for designing new and more effective drugs. Due to the still high costs and to the several limitations of experimental techniques, it is most often desirable to predict these ligand-protein complexes in silico, particularly when screening for new putative drugs from databases of millions of compounds. While virtual screening based on molecular docking is widely used for this purpose, it generally fails in mimicking binding events associated with large conformational changes in the protein, particularly when the latter involve multiple domains. In this work, we describe a new methodology aimed at generating bound-like conformations of very flexible and allosteric proteins bearing multiple binding sites. Validation was performed on the enzyme adenylate kinase (ADK), a paradigmatic example of proteins that undergo very large conformational changes upon ligand binding. By only exploiting the unbound structure and the putative binding sites of the protein, we generated a significant fraction of bound-like structures, which employed in ensemble-docking calculations allowed to find native-like poses of substrates, inhibitors, and catalytically incompetent binders. Our protocol provides a general framework for the generation of bound-like conformations of flexible proteins that are suitable to host different ligands, demonstrating high sensitivity to the fine chemical details that regulate protein's activity. We foresee applications in virtual screening for difficult targets, prediction of the impact of amino acid mutations on structure and dynamics, and protein engineering.