title: Experimental Correlation of Substrate Position with Reaction Outcome in the Aliphatic Halogenase, SyrB2.
abstract: The iron(II)- and 2-(oxo)glutarate-dependent (Fe/2OG) oxygenases catalyze an array of challenging transformations, but how individual members of the enzyme family direct different outcomes is poorly understood. The Fe/2OG halogenase, SyrB2, chlorinates C4 of its native substrate, l-threonine appended to the carrier protein, SyrB1, but hydroxylates C5 of l-norvaline and, to a lesser extent, C4 of l-aminobutyric acid when SyrB1 presents these non-native amino acids. To test the hypothesis that positioning of the targeted carbon dictates the outcome, we defined the positions of these three substrates by measuring hyperfine couplings between substrate deuterium atoms and the stable, EPR-active iron-nitrosyl adduct, a surrogate for reaction intermediates. The Fe-(2)H distances and N-Fe-(2)H angles, which vary from 4.2 Å and 85° for threonine to 3.4 Å and 65° for norvaline, rationalize the trends in reactivity. This experimental correlation of position to outcome should aid in judging from structural data on other Fe/2OG enzymes whether they suppress hydroxylation or form hydroxylated intermediates on the pathways to other outcomes.
authors.affiliation: Departments of †Chemistry and of ‡Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania 16802, United States.
authors.forename: Ryan J
authors.initials: RJ
authors.lastname: Martinie
authors.affiliation: Departments of †Chemistry and of ‡Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania 16802, United States.
authors.forename: Jovan
authors.initials: J
authors.lastname: Livada
authors.affiliation: Departments of †Chemistry and of ‡Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania 16802, United States.
authors.forename: Wei-chen
authors.initials: WC
authors.lastname: Chang
authors.affiliation: Departments of †Chemistry and of ‡Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania 16802, United States.
authors.forename: Michael T
authors.initials: MT
authors.lastname: Green
authors.affiliation: Departments of †Chemistry and of ‡Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania 16802, United States.
authors.forename: Carsten
authors.initials: C
authors.lastname: Krebs
authors.affiliation: Departments of †Chemistry and of ‡Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania 16802, United States.
authors.forename: J Martin
authors.initials: JM
authors.lastname: Bollinger
authors.affiliation: Departments of †Chemistry and of ‡Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania 16802, United States.
authors.forename: Alexey
authors.initials: A
authors.lastname: Silakov
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D008968:Molecular Conformation; D044224:Pseudomonas syringae / Q000201:enzymology*; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins