title: A common late-stage intermediate in catalysis by 2-hydroxyethyl-phosphonate dioxygenase and methylphosphonate synthase.
abstract: 2-Hydroxyethylphosphonate dioxygenase (HEPD) and methylphosphonate synthase (MPnS) are nonheme iron oxygenases that both catalyze the carbon-carbon bond cleavage of 2-hydroxyethylphosphonate but generate different products. Substrate labeling experiments led to a mechanistic hypothesis in which the fate of a common intermediate determined product identity. We report here the generation of a bifunctional mutant of HEPD (E176H) that exhibits the activity of both HEPD and MPnS. The product distribution of the mutant is sensitive to a substrate isotope effect, consistent with an isotope-sensitive branching mechanism involving a common intermediate. The X-ray structure of the mutant was determined and suggested that the introduced histidine does not coordinate the active site metal, unlike the iron-binding glutamate it replaced.
authors.affiliation: Howard Hughes Medical Institute and Department of Chemistry, University of Illinois at Urbana-Champaign , 600 South Mathews Avenue, Urbana, Illinois 61801, United States.
authors.forename: Spencer C
authors.initials: SC
authors.lastname: Peck
authors.forename: Jonathan R
authors.initials: JR
authors.lastname: Chekan
authors.forename: Emily C
authors.initials: EC
authors.lastname: Ulrich
authors.forename: Satish K
authors.initials: SK
authors.lastname: Nair
authors.forename: Wilfred A
authors.initials: WA
authors.lastname: van der Donk
mesh_terms: D002384:Catalysis; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D049308:Dioxygenases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018698:Glutamic Acid / Q000235:genetics / Q000378:metabolism; D006639:Histidine; D009682:Magnetic Resonance Spectroscopy; D009154:Mutation; D063065:Organophosphonates / Q000378:metabolism; D009943:Organophosphorus Compounds / Q000378:metabolism; D010105:Oxygenases / Q000737:chemistry / Q000378:metabolism*; D011487:Protein Conformation
chemical_list: C519566:2-hydroxyethyl phosphonate; D063065:Organophosphonates; D009943:Organophosphorus Compounds; C032627:methylphosphonic acid; D018698:Glutamic Acid; D006639:Histidine; D010105:Oxygenases; D049308:Dioxygenases; C577763:methylphosphonate synthase, Nitrosopumilus maritimus