title: Structural analysis of the zinc hydroxide-Thr-199-Glu-106 hydrogen-bond network in human carbonic anhydrase II.
abstract: The significance of the zinc hydroxide-Thr-199-Glu-106 hydrogen-bond network in the active site of human carbonic anhydrase II has been examined by X-ray crystallographic analyses of site-specific mutants. Mutants with Ala-199 and Ala-106 or Gln-106 have low catalytic activities, while a mutant with Asp-106 has almost full CO2 hydration activity. The structures of these four mutants, as well as that of the bicarbonate complex of the mutant with Ala-199, have been determined at 1.7 to 2.2 A resolution. Removal of the gamma atoms of residue 199 leads to a distorted tetrahedral geometry at the zinc ion, and a catalytically important zinc-bound water molecule has moved towards Glu-106. In the bicarbonate complex of the mutant with Ala-199 one oxygen atom from bicarbonate binds to zinc without displacing this water molecule. Tetrahedral coordination geometries are retained in the mutants at position 106. The mutants with Ala-106 and Gln-106 have a zinc-bound sulfate ion, whereas this sulfate site is only partially occupied in the mutant with Asp-106. The hydrogen-bond network seems to be "reversed" in the mutants with Ala-106 and Gln-106. The network is preserved as in native enzyme in the mutant with Asp-106 but the side chain of Asp-106 is more extended than that of Glu-106 in the native enzyme. These results illustrate the importance of Glu-106 and Thr-199 for controlling the precise coordination geometry of the zinc ion and its ligand preferences which results in an optimal orientation of a zinc-bound hydroxide ion for an attack on the CO2 substrate.
authors.affiliation: Department of Biochemistry, University of Umeå, Sweden.
authors.forename: Y
authors.initials: Y
authors.lastname: Xue
authors.forename: A
authors.initials: A
authors.lastname: Liljas
authors.forename: B H
authors.initials: BH
authors.lastname: Jonsson
authors.forename: S
authors.initials: S
authors.lastname: Lindskog
mesh_terms: D002256:Carbonic Anhydrases / Q000737:chemistry* / Q000378:metabolism; D002384:Catalysis; D003198:Computer Simulation; D018360:Crystallography, X-Ray; D005971:Glutamates / Q000737:chemistry; D018698:Glutamic Acid; D006801:Humans; D006860:Hydrogen Bonding; D006878:Hydroxides / Q000737:chemistry* / Q000378:metabolism; D008956:Models, Chemical; D008958:Models, Molecular; D016296:Mutagenesis; D011485:Protein Binding; D013912:Threonine / Q000737:chemistry; D017967:Zinc Compounds / Q000737:chemistry* / Q000378:metabolism
chemical_list: D005971:Glutamates; D006878:Hydroxides; D017967:Zinc Compounds; D013912:Threonine; D018698:Glutamic Acid; D002256:Carbonic Anhydrases; C052745:zinc hydroxide