Effect of Asp122 Mutation on the Hydride Transfer in <i>E. coli</i> DHFR Demonstrates the Goldilocks of Enzyme Flexibility

“…Indeed, the normalized B ‐factors of residues 167–170 are elevated in P169S, implying increased conformational flexibility in the second sphere around AASAL. It is well known that changes in active site and second sphere flexibility and dynamics are linked to changes in catalysis in a number of enzymes , including cyclophilin A , dihydrofolate reductase , pentaerythritol tetranitrate reductase , and liver alcohol dehydrogenase . The latter three of these four examples all catalyze a hydride transfer step, which is highlighted as the aspect of the catalytic mechanism that is affected by changes in protein dynamics and flexibility.…”

“…Supplementary File 1c: Fisher Exact Test p-values for the null hypothesis that conserved positions and inactivating mutants are independent. Calculations were made over a range of conservation definitions chosen to result in an equal number positions as the sector positions in Supplementary File 1b (23,36,40,and 49 positions respectively. In all cases, the null hypothesis is rejected at a confidence level of 0.05 or better (red), and inactivating mutations are enriched at conserved positions beyond expectation due to random chance.…”

“…In the absence of a proper computational model, it is almost impossible to assess the dielectric effects in the protein and thus to estimate the strength of electrostatic effects. However, pioneering work [16] provided the first quantitative hint that electrostatic effects may have a major role in enzyme catalysis, suggesting that the catalytic power of enzymes is almost exclusively due to electrostatic effects and specifically to the preorganized electrostatic environment of their active sites [15] , [17] . Since all quantum chemistry is electrostatic ,“electrostatic catalysis” [16] takes into account the effects of the protein charges, permanent/induced dipoles, metal ions and the solvation by bound water molecules [15] .…”