title: Structure of the triosephosphate isomerase-phosphoglycolohydroxamate complex: an analogue of the intermediate on the reaction pathway.
abstract: The glycolytic enzyme triosephosphate isomerase (TIM) catalyzes the interconversion of the three-carbon sugars dihydroxyacetone phosphate (DHAP) and D-glyceraldehyde 3-phosphate (GAP) at a rate limited by the diffusion of substrate to the enzyme. We have solved the three-dimensional structure of TIM complexed with a reactive intermediate analogue, phosphoglycolohydroxamate (PGH), at 1.9-A resolution and have refined the structure to an R-factor of 18%. Analysis of the refined structure reveals the geometry of the active-site residues and the interactions they make with the inhibitor and, by analogy, the substrates. The structure is consistent with an acid-base mechanism in which the carboxylate of Glu-165 abstracts a proton from carbon while His-95 donates a proton to oxygen to form an enediol (or enediolate) intermediate. The conformation of the bound substrate stereoelectronically favors proton transfer from substrate carbon to the syn orbital of Glu-165. The crystal structure suggests that His-95 is neutral rather than cationic in the ground state and therefore would have to function as an imidazole acid instead of the usual imidazolium. Lys-12 is oriented so as to polarize the substrate oxygens by hydrogen bonding and/or electrostatic interaction, providing stabilization for the charged transition state. Asn-10 may play a similar role.
authors.affiliation: Department of Chemistry, Massachusetts Institute of Technology, Cambridge 02139.
authors.forename: R C
authors.initials: RC
authors.lastname: Davenport
authors.forename: P A
authors.initials: PA
authors.lastname: Bash
authors.forename: B A
authors.initials: BA
authors.lastname: Seaton
authors.forename: M
authors.initials: M
authors.lastname: Karplus
authors.forename: G A
authors.initials: GA
authors.lastname: Petsko
authors.forename: D
authors.initials: D
authors.lastname: Ringe
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D006639:Histidine; D006877:Hydroxamic Acids / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D008968:Molecular Conformation; D015394:Molecular Structure; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae; D014305:Triose-Phosphate Isomerase / Q000737:chemistry* / Q000378:metabolism; D014961:X-Ray Diffraction
chemical_list: D006877:Hydroxamic Acids; D006639:Histidine; C008886:phosphoglycolohydroxamate; D014305:Triose-Phosphate Isomerase