3.8.1 MM/GBSA free energy calculations
To understand the thermodynamics behind the protein-DNA complexes,
MM-GBSA was calculated to determine the binding mechanisms. The
calculations were performed to calculate the binding energies of WT
protein comparing it to mutant protein in the DNA-protein complex. Based
on the 200ns MD trajectories of WT and variants the energy analysis and
its corresponding component as calculated and reported in Table 3. The
results indicated high binding energy (-216.33 Kcal/mol) compared to
most other variants. The point mutations decreased the positive polar
term resulting in overall increase of the negative term promoting
DNA-protein complex formation. The energy components included here are
Van der Waals, electrostatic interaction, non-polar and polar energies
contributing to the total binding energy and favouring complex
formation. Primarily, the stability of the complex was majorly due to
the electrostatic interaction energy whereas other energy terms
contributed very less to the total energy of the complex. The
spontaneity of this interactions is highlighted by the high negative
value of the electrostatic interactions, whilst polar solvation energies
hinders the interactions between DNA and protein. Interestingly, some of
the residues showed higher binding stability when mutated resulting in
higher negative values in T144R, A149V and Q188E. N-terminal R
(arginine) showed a loss in interaction energy (-188.74 Kcal/mol) when
mutated to lysine. However, when mutated to tryptophan, it showed
considerable increase in binding energy (-224.41) of the mutant. Studies
have highlighted the role of tryptophan in DNA binding domain to be
important, which might be the case in this mutant also76. The same effect was
observed on the C-terminal R when mutated to tryptophan (R198W= -224.43
Kcal/mol). But when mutated to cysteine (-186.36 Kcal/mol) and histidine
(-185.16 Kcal/mol) there was a loss of interaction energy of the mutant.
These residues were also found to be highly conserved in the Consurf
analysis. Furthermore, this also implies about the role of these
conserved residues to predominantly be the fundamental residues driving
a compact interaction with DNA.