title: Spectroscopic and computational studies of α-keto acid binding to Dke1: understanding the role of the facial triad and the reactivity of β-diketones.
abstract: The O(2) activating mononuclear nonheme iron enzymes generally have a common facial triad (two histidine and one carboxylate (Asp or Glu) residue) ligating Fe(II) at the active site. Exceptions to this motif have recently been identified in nonheme enzymes, including a 3His triad in the diketone cleaving dioxygenase Dke1. This enzyme is used to explore the role of the facial triad in directing reactivity. A combination of spectroscopic studies (UV-vis absorption, MCD, and resonance Raman) and DFT calculations is used to define the nature of the binding of the α-keto acid, 4-hydroxyphenlpyruvate (HPP), to the active site in Dke1 and the origin of the atypical cleavage (C2-C3 instead of C1-C2) pattern exhibited by this enzyme in the reaction of α-keto acids with dioxygen. The reduced charge of the 3His triad induces α-keto acid binding as the enolate dianion, rather than the keto monoanion, found for α-keto acid binding to the 2His/1 carboxylate facial triad enzymes. The mechanistic insight from the reactivity of Dke1 with the α-keto acid substrate is then extended to understand the reaction mechanism of this enzyme with its native substrate, acac. This study defines a key role for the 2His/1 carboxylate facial triad in α-keto acid-dependent mononuclear nonheme iron enzymes in stabilizing the bound α-keto acid as a monoanion for its decarboxylation to provide the two additional electrons required for O(2) activation.
authors.affiliation: Department of Chemistry, Stanford University, Stanford, California 94305, United States.
authors.forename: Adrienne R
authors.initials: AR
authors.lastname: Diebold
authors.forename: Grit D
authors.initials: GD
authors.lastname: Straganz
authors.forename: Edward I
authors.initials: EI
authors.lastname: Solomon
mesh_terms: D000150:Acinetobacter / Q000737:chemistry / Q000201:enzymology* / Q000378:metabolism; D001665:Binding Sites; D049308:Dioxygenases / Q000737:chemistry / Q000378:metabolism*; D007651:Keto Acids / Q000378:metabolism*; D007659:Ketones / Q000378:metabolism; D008958:Models, Molecular; D010100:Oxygen / Q000378:metabolism*; D011485:Protein Binding; D013057:Spectrum Analysis
chemical_list: D007651:Keto Acids; D007659:Ketones; D049308:Dioxygenases; D010100:Oxygen