@Article{IPB-361, author = {Meng, S. and Ji, Y. and Liu, L. and Davari, M. D. and Schwaneberg, U.}, title = {{Modulating the coupling efficiency of P450 BM3 by controlling water diffusion through access tunnel engineering}}, year = {2022}, pages = {e202102434}, journal = {ChemSusChem}, doi = {10.1002/cssc.202102434}, url = {https://doi.org/10.1002/cssc.202102434}, volume = {15}, abstract = {Cytochrome P450s has gained the great interest for their broad  substrate  scope  in the catalysis  of  oxidation  reactions  for pharmaceuticals,  plastics,  and  hormones.  However, achieving  high coupling efficiency by the engineering of P450s is still a big challenge.The presence of extra water around the active site is deemed to be related touncoupling.  Herein,  we  engineered  the  access  tunnels  of P450 BM3 from Bacillus megaterium to control water access from bulk solvent  to the active  site.  Nine  residues  located  in  tunnels  were investigated bysite  saturationmutagenesisto reduce  the  water diffusion,therefore,improvingthe  coupling  efficiency.  Finally,  the recombinedvariant N319L/T411V/T436A showed improved coupling efficiency (from 31.2%  to 52.6%). Tunnel  polarity  analysis  and  MD simulation  further  proved  that  reduced  water moleculesaround the active  site  lead  to  higher  coupling  efficiency. Overall,  our  research provides   valuable   insight   on   improved   coupling   efficiency   by controlling water diffusion through tunnel engineering.} }