The work performed within the UBioRec project was divided into different work packages.In the first step, the objectives were to test and improve force fields parameters. Special attention has been considered for the quality of the atomic charges and torsional potentials, as they are expected to be crucial for the energetics of ligand binding/unbinding to/from the binding sites of targets selected. The use of a99SB-disp force field for the targets was corroborated as the best option in comparison with different force fields realized along the first semester of the project.
Then, the main work package involve the testing and developing of different enhanced-sampling algorithms for the calculation of binding free energies, with a good balance between accuracy, computational cost and speed. The main algorithms used were based on funnel-shaped restraint metadynamics. The advantage of using funnel-shaped restraints is the increase it provides to the speed of convergence of the simulations, and favouring re-crossings between bound and unbound states. Additionally, the general CVs that define the funnel restraints avoid extra simulations, reducing the number of computational resources needed for more computational demanding CVs. The combination of fun-metaD with SWISH (Sampling Water Interfaces through Scaled Hamiltonians), a Hamiltonian replica-exchange based method recently developed by Gervasio’s group was also used.During the UBioRec project, a novel application of the method was applied helping to understand the hydration/dehydration process during the dynamics itself in an unprecedented manner. An additional crucial idea developed in the UBioRec project is the definition of general convergence criteria, which help us to objectively evaluate the convergence of enhanced sampling simulations, independently of the experimental binding affinities available. The criteria established are general and have been applied to all the methods mentioned above. From these results, I published two papers, one as a first author and one as corresponding author.
Additional steps were i) the combination of computational approaches at different scales, and ii) obtaining the experimental data by employing biophysical techniques. For the first point, the results from the previous ES simulations were used to get the most relevant clusters for the QM/MM re-weighting calculations. For the latest point, the expression and purification of WT and the mutated targets were successfully obtained. SPR experiments were run providing information about binding kinetics and affinity. These results were needed to complement and validate the theoretical results obtained during the developing of the project. Finally, for the creation of reference data and platform all my efforts were concentrated on the compilation of the data obtained during the UBioRec project, together with the scripts and inputs used during the simulation process. All this information was structured in a GitLab repository in order to organize and develop the platform. All the files required to reproduce the simulations have been uploaded to the Plumed-Nest repository. The source code for the new CVs developed, are now part of the development branch of the Plumed plugin and will be made available in the next releases for the whole community.
