Computational Alanine Scanning MutagenesisAn Improved Methodological Approach for Protein–DNA Complexes

Proteins
and protein-based complexes are the basis of many key
systems in nature and have been the subject of intense research in
the last decades, in an attempt to acquire comprehensive knowledge
of reactions that take place in nature. Computational Alanine Scanning
Mutagenesis approaches have been extensively used in the study of
protein interfaces and in the determination of the most important
residues for complex formation, the Hot-spots. However, as it is usually
applied to the study of protein–protein interfaces, we tried
to modify and apply it to the study of protein–DNA interfaces,
which are also crucial in nature but have not been the subject of
as much research. In this work, we carry out MD simulations of seven
protein–DNA complexes and tested the influence of the variation
of different parameters on the determination of the binding free energy
terms (ΔΔGbinding) of 78 mutations:
solvent representation, internal dielectric constant, Linear and Nonlinear
Poisson–Boltzmann equation, Generalized Born model, simulation
time, number of structures analyzed, number of MD trajectories, force
field used, and energetic terms involved. Overall, this new approach
gave an average error of 1.55 kcal/mol, and P, R, F1, accuracy, and specificity values of 0.78, 0.50, 0.61,
0.77, and 0.92, respectively. This improved computational alanine
scanning mutagenesis approach may serve as a tool to explore the behavior
of this important class of complexes.