title: Structures of horse liver alcohol dehydrogenase complexed with NAD+ and substituted benzyl alcohols.
abstract: Structures of the enzyme complexed with NAD+ and 2,3,4,5,6-pentafluorobenzyl alcohol were determined by X-ray crystallography at a resolution of 2.1 A and to a refinement R value of 18.3% for a monoclinic (P2(1)) form and to 2.4 A and an R value of 18.9% for a triclinic crystal form. The pentafluorobenzyl alcohol does not react, due to electron withdrawal by the fluorine atoms. A structure with NAD+ and p-bromobenzyl alcohol in the monoclinic form was also determined at 2.5 A and an R value of 16.7%. The conformations of the subunits in the monoclinic and triclinic crystal forms are very similar. The dimer is the asymmetric unit, and a rigid body rotation closes the cleft between the coenzyme and catalytic domains upon complex formation. In the monoclinic form, this conformational change is described by a rotation of 9 degrees in one subunit and 10 degrees in the other. The pentafluoro- and p-bromobenzyl alcohols bind in overlapping positions. The hydroxyl group of each alcohol is ligated to the catalytic zinc and participates in an extensive hydrogen-bonded network that includes the imidazole group of His-51, which can act as a base and shuttle a proton to solvent. The hydroxymethyl carbon of the pentafluorobenzyl alcohol is 3.4 A from C4 of the nicotinamide ring, and the pro-R hydrogen is in a good position for direct transfer to C4. The p-bromobenzyl alcohol may react after small rotations around single bonds of the alcohol. These structures should approximate the active Michaelis-Menten complexes.
authors.affiliation: Department of Molecular Biology, Swedish University of Agricultural Sciences, Uppsala.
authors.forename: S
authors.initials: S
authors.lastname: Ramaswamy
authors.forename: H
authors.initials: H
authors.lastname: Eklund
authors.forename: B V
authors.initials: BV
authors.lastname: Plapp
mesh_terms: D000426:Alcohol Dehydrogenase / Q000737:chemistry* / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001592:Benzyl Alcohols / Q000378:metabolism*; D001665:Binding Sites; D018360:Crystallography, X-Ray / Q000379:methods; D005583:Fourier Analysis; D006639:Histidine; D006736:Horses; D006860:Hydrogen Bonding; D008099:Liver / Q000201:enzymology*; D046911:Macromolecular Substances; D009243:NAD / Q000378:metabolism*; D011487:Protein Conformation; D013816:Thermodynamics; D005464:Fluorobenzenes; D005466:Fluorocarbons
chemical_list: D000426:Alcohol Dehydrogenase; D001592:Benzyl Alcohols; D006639:Histidine; D046911:Macromolecular Substances; D009243:NAD; C087170:2,3,4,5,6-pentafluorobenzyl alcohol; D005464:Fluorobenzenes; D005466:Fluorocarbons