title: Perturbation of the Conformational Dynamics of an Active-Site Loop Alters Enzyme Activity.
abstract: The role of internal dynamics in enzyme function is highly debated. Specifically, how small changes in structure far away from the reaction site alter protein dynamics and overall enzyme mechanisms is of wide interest in protein engineering. Using RNase A as a model, we demonstrate that elimination of a single methyl group located >10 Å away from the reaction site significantly alters conformational integrity and binding properties of the enzyme. This A109G mutation does not perturb structure or thermodynamic stability, both in the apo and ligand-bound states. However, significant enhancement in conformational dynamics was observed for the bound variant, as probed over nano- to millisecond timescales, resulting in major ligand repositioning. These results illustrate the large effects caused by small changes in structure on long-range conformational dynamics and ligand specificities within proteins, further supporting the importance of preserving wild-type dynamics in enzyme systems that rely on flexibility for function.
authors.affiliation: INRS-Institut Armand-Frappier, Université du Québec, 531 Boulevard des Prairies, Laval, QC H7V 1B7, Canada.
authors.forename: Donald
authors.initials: D
authors.lastname: Gagné
authors.affiliation: Department of Biochemistry and Molecular Genetics, University of Illinois at Chicago, 900 South Ashland, Chicago, IL 60607, USA.
authors.forename: Rachel L
authors.initials: RL
authors.lastname: French
authors.affiliation: INRS-Institut Armand-Frappier, Université du Québec, 531 Boulevard des Prairies, Laval, QC H7V 1B7, Canada.
authors.forename: Chitra
authors.initials: C
authors.lastname: Narayanan
authors.affiliation: Department of Biochemistry and Molecular Genetics, University of Illinois at Chicago, 900 South Ashland, Chicago, IL 60607, USA.
authors.forename: Miljan
authors.initials: M
authors.lastname: Simonović
authors.affiliation: Computational Biology Institute and Computer Science and Mathematics Division, Oak Ridge National Laboratory, 1 Bethel Valley Road, Oak Ridge, TN 37830, USA; Department of Biochemistry, Cellular and Molecular Biology, University of Tennessee, Knoxville, TN 37996, USA.
authors.forename: Pratul K
authors.initials: PK
authors.lastname: Agarwal
authors.affiliation: INRS-Institut Armand-Frappier, Université du Québec, 531 Boulevard des Prairies, Laval, QC H7V 1B7, Canada; PROTEO, the Québec Network for Research on Protein Function, Engineering, and Applications, 1045 Avenue de la Médecine, Université Laval, QC G1V 0A6, Canada; GRASP, the Groupe de Recherche Axé sur la Structure des Protéines, 3649 Promenade Sir William Osler, McGill University, Montréal, QC H3G 0B1, Canada. Electronic address: nicolas.doucet@iaf.inrs.ca.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Doucet
mesh_terms: D000249:Adenosine Monophosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D020134:Catalytic Domain; D002417:Cattle; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D012259:Ribonuclease, Pancreatic / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014542:Uridine Monophosphate / Q000378:metabolism
chemical_list: D000249:Adenosine Monophosphate; D014542:Uridine Monophosphate; D012259:Ribonuclease, Pancreatic