title: Unravelling the Molecular Origin of the Regiospecificity in Extradiol Catechol Dioxygenases.
abstract: Many factors have been suggested to control the selectivity for extradiol or intradiol cleavage in catechol dioxygenases. The varied selectivity of model complexes and the ability to force an extradiol enzyme to do intradiol cleavage indicate that the problem may be complex. In this paper we focus on the regiospecificity of the proximal extradiol dioxygenase, homoprotocatechuate 2,3-dioxygenase (HPCD), for which considerable advances have been made in our understanding of the mechanism from an experimental and computational standpoint. Two key steps in the reaction mechanism were investigated: (1) attack of the substrate by the superoxide moiety and (2) attack of the substrate by the oxyl radical generated by O-O bond cleavage. The selectivity at both steps was investigated through a systematic study of the role of the substrate and the first and second coordination spheres. For the isolated native substrate, intradiol cleavage is calculated to be both kinetically and thermodynamically favored, therefore nature must use the enzyme environment to reverse this preference. Two second sphere residues were found to play key roles in controlling the regiospecificity of the reaction: Tyr257 and His200. Tyr257 controls the selectivity by modulating the electronic structure of the substrate, while His200 controls selectivity through steric effects and by preventing alternative pathways to intradiol cleavage.
authors.affiliation: Max-Planck Institute for Chemical Energy Conversion , Stiftstrasse 34-36, D-45470 Mülheim an der Ruhr, Germany.|Avondale College of Higher Education , Cooranbong, New South Wales 2265, Australia.
authors.forename: Gemma J
authors.initials: GJ
authors.lastname: Christian
authors.affiliation: Max-Planck Institute for Chemical Energy Conversion , Stiftstrasse 34-36, D-45470 Mülheim an der Ruhr, Germany.
authors.forename: Frank
authors.initials: F
authors.lastname: Neese
authors.affiliation: Max-Planck Institute for Chemical Energy Conversion , Stiftstrasse 34-36, D-45470 Mülheim an der Ruhr, Germany.
authors.forename: Shengfa
authors.initials: S
authors.lastname: Ye
mesh_terms: D055162:Biocatalysis; D002396:Catechols / Q000737:chemistry / Q000378:metabolism*; D015394:Molecular Structure; D010105:Oxygenases / Q000737:chemistry* / Q000378:metabolism*; D011789:Quantum Theory; D013237:Stereoisomerism
chemical_list: D002396:Catechols; D010105:Oxygenases; C439587:extradiol dioxygenase