title: Insights into the Thiamine Diphosphate Enzyme Activation Mechanism: Computational Model for Transketolase Using a Quantum Mechanical/Molecular Mechanical Method.
abstract: We propose the first computational model for transketolase (TK), a thiamine diphosphate (ThDP)-dependent enzyme, using a quantum mechanical/molecular mechanical method on the basis of crystallographic TK structures from yeast and Escherichia coli, together with experimental kinetic data reported in the literature with wild-type and mutant TK. This model allowed us to define a new route for ThDP activation in the enzyme environment. We evidenced a strong interaction between ThDP and Glu418B of the TK active site, itself stabilized by Glu162A. The crucial point highlighted here is that deprotonation of ThDP C2 is not performed by ThDP N4' as reported in the literature, but by His481B, involving a HOH688A molecule bridge. Thus, ThDP N4' is converted from an amino form to an iminium form, ensuring the stabilization of the C2 carbanion or carbene. Finally, ThDP activation proceeds via an intermolecular process and not by an intramolecular one as reported in the literature. More generally, this proposed ThDP activation mechanism can be applied to some other ThDP-dependent enzymes and used to define the entire TK mechanism with donor and acceptor substrates more accurately.
authors.affiliation: Université Clermont Auvergne, Université Blaise-Pascal , Institut de Chimie de Clermont-Ferrand, BP 10448, F-63000 Clermont-Ferrand, France.|CNRS , UMR 6296, ICCF, F-63178 Aubiere, France.
authors.forename: Lionel
authors.initials: L
authors.lastname: Nauton
authors.affiliation: Université Clermont Auvergne, Université Blaise-Pascal , Institut de Chimie de Clermont-Ferrand, BP 10448, F-63000 Clermont-Ferrand, France.|CNRS , UMR 6296, ICCF, F-63178 Aubiere, France.
authors.forename: Virgil
authors.initials: V
authors.lastname: Hélaine
authors.affiliation: Université Clermont Auvergne, Université Blaise-Pascal , Institut de Chimie de Clermont-Ferrand, BP 10448, F-63000 Clermont-Ferrand, France.|CNRS , UMR 6296, ICCF, F-63178 Aubiere, France.
authors.forename: Vincent
authors.initials: V
authors.lastname: Théry
authors.affiliation: Université Clermont Auvergne, Université Blaise-Pascal , Institut de Chimie de Clermont-Ferrand, BP 10448, F-63000 Clermont-Ferrand, France.|CNRS , UMR 6296, ICCF, F-63178 Aubiere, France.
authors.forename: Laurence
authors.initials: L
authors.lastname: Hecquet
mesh_terms: D019943:Amino Acid Substitution; D001665:Binding Sites; D055162:Biocatalysis; D020134:Catalytic Domain; D019295:Computational Biology; D030562:Databases, Protein; D019281:Dimerization; D004735:Energy Transfer; D004789:Enzyme Activation; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008968:Molecular Conformation; D056004:Molecular Dynamics Simulation; D009154:Mutation; D011789:Quantum Theory; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D040681:Structural Homology, Protein; D013816:Thermodynamics; D013835:Thiamine Pyrophosphate / Q000737:chemistry* / Q000378:metabolism; D014174:Transketolase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014803:Vitamin B Complex / Q000737:chemistry* / Q000378:metabolism
chemical_list: D029968:Escherichia coli Proteins; D029701:Saccharomyces cerevisiae Proteins; D014803:Vitamin B Complex; C084088:TKL1 protein, S cerevisiae; D014174:Transketolase; D013835:Thiamine Pyrophosphate