Functionally important conformations of the Met20 loop in dihydrofolate reductase are populated by rapid thermal fluctuations.
Conformational changes in enzymes are well recognized to play an important role in the organization of the reactive groups for efficient catalysis. This study reveals atomic and energetic details of the conformational change process that precedes the catalytic reaction of the enzyme dihydrofolate reductase. The computed free energy profile provides insights into the ligand binding mechanism and a quantitative estimate of barrier heights separating different conformational states along the pathway. Studies show that the ternary complex comprised of NADPH cofactor and substrate dihydrofolate undergoes transitions between a closed state and an occluded state via an intermediate "open" conformation. During these transitions the largest conformational change occurs in the Met20 loop of DHFR and is accompanied by the motion of the cofactor into and out of the binding pocket. When the cofactor is out of the binding pocket, the enzyme frequently samples open and occluded conformations with a small (approximately 5 k(B)T) free energy barrier between the two states. However, when the cofactor is in the binding pocket, the closed conformation is thermodynamically most favored. The determination of a profile characterizing the position-dependent diffusion of the Met20 loop allowed us to apply reaction rate theory and deduce the kinetics of loop motions based on the computed free energy landscape.
Arora
Karunesh
K
Department of Chemistry and Biophysics Program, University of Michigan, 930 North University Avenue, Ann Arbor, Michigan 48109, USA.
Brooks Iii
Charles L
CL