title: Evidence that oxidative dephosphorylation by the nonheme Fe(II), α-ketoglutarate:UMP oxygenase occurs by stereospecific hydroxylation.
abstract: LipL and Cpr19 are nonheme, mononuclear Fe(II)-dependent, α-ketoglutarate (αKG):UMP oxygenases that catalyze the formation of CO2, succinate, phosphate, and uridine-5'-aldehyde, the last of which is a biosynthetic precursor for several nucleoside antibiotics that inhibit bacterial translocase I (MraY). To better understand the chemistry underlying this unusual oxidative dephosphorylation and establish a mechanistic framework for LipL and Cpr19, we report herein the synthesis of two biochemical probes-[1',3',4',5',5'-2H]UMP and the phosphonate derivative of UMP-and their activity with both enzymes. The results are consistent with a reaction coordinate that proceeds through the loss of one2H atom of [1',3',4',5',5'-2H]UMP and stereospecific hydroxylation geminal to the phosphoester to form a cryptic intermediate, (5'R)-5'-hydroxy-UMP. Thus, these enzyme catalysts can additionally be assigned as UMP hydroxylase-phospholyases.
authors.affiliation: Department of Pharmaceutical Sciences, University of Kentucky, Lexington, KY, USA.
authors.forename: Anwesha
authors.initials: A
authors.lastname: Goswami
authors.affiliation: Department of Pharmaceutical Sciences, University of Kentucky, Lexington, KY, USA.
authors.forename: Xiaodong
authors.initials: X
authors.lastname: Liu
authors.affiliation: Department of Pharmaceutical Sciences, University of Kentucky, Lexington, KY, USA.
authors.forename: Wenlong
authors.initials: W
authors.lastname: Cai
authors.affiliation: Department of Pharmaceutical Sciences, University of Wisconsin-Madison, WI, USA.
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Wyche
authors.affiliation: Department of Pharmaceutical Sciences, University of Wisconsin-Madison, WI, USA.
authors.forename: Tim S
authors.initials: TS
authors.lastname: Bugni
authors.affiliation: Nucleosides and Phosphorylated Effectors Team, IBMM, UMR5247 CNRS University Montpellier, France.
authors.forename: Maïa
authors.initials: M
authors.lastname: Meurillon
authors.affiliation: Nucleosides and Phosphorylated Effectors Team, IBMM, UMR5247 CNRS University Montpellier, France.
authors.forename: Suzanne
authors.initials: S
authors.lastname: Peyrottes
authors.affiliation: Nucleosides and Phosphorylated Effectors Team, IBMM, UMR5247 CNRS University Montpellier, France.
authors.forename: Christian
authors.initials: C
authors.lastname: Perigaud
authors.affiliation: Biologics Technology Research Laboratories, R&D Division, Daiichi Sankyo Co., Ltd., Gunma, Japan.
authors.forename: Koichi
authors.initials: K
authors.lastname: Nonaka
authors.affiliation: Department of Pharmaceutical Sciences, University of Kentucky, Lexington, KY, USA.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Rohr
authors.affiliation: Department of Pharmaceutical Sciences, University of Kentucky, Lexington, KY, USA.
authors.forename: Steven G
authors.initials: SG
authors.lastname: Van Lanen
mesh_terms: D055162:Biocatalysis; D006418:Heme / Q000378:metabolism*; D006859:Hydrogen / Q000378:metabolism; D006900:Hydroxylation; D007501:Iron / Q000378:metabolism*; D007656:Ketoglutaric Acids / Q000378:metabolism*; D010084:Oxidation-Reduction; D010105:Oxygenases / Q000378:metabolism*; D010766:Phosphorylation; D013237:Stereoisomerism; D013379:Substrate Specificity; D014542:Uridine Monophosphate / Q000737:chemistry / Q000378:metabolism*
chemical_list: D007656:Ketoglutaric Acids; D006418:Heme; D006859:Hydrogen; D007501:Iron; D014542:Uridine Monophosphate; D010105:Oxygenases
keywords: antibiotic; biosynthesis; nonheme iron; nucleoside; oxygenase; translocase I