title: Selective hydroxylation of alkanes by an extracellular fungal peroxygenase.
abstract: Fungal peroxygenases are novel extracellular heme-thiolate biocatalysts that are capable of catalyzing the selective monooxygenation of diverse organic compounds, using only H(2)O(2) as a cosubstrate. Little is known about the physiological role or the catalytic mechanism of these enzymes. We have found that the peroxygenase secreted by Agrocybe aegerita catalyzes the H(2)O(2)-dependent hydroxylation of linear alkanes at the 2-position and 3-position with high efficiency, as well as the regioselective monooxygenation of branched and cyclic alkanes. Experiments with n-heptane and n-octane showed that the hydroxylation proceeded with complete stereoselectivity for the (R)-enantiomer of the corresponding 3-alcohol. Investigations with a number of model substrates provided information about the route of alkane hydroxylation: (a) the hydroxylation of cyclohexane mediated by H(2)(18)(2) resulted in complete incorporation of (18)O into the hydroxyl group of the product cyclohexanol; (b) the hydroxylation of n-hexane-1,1,1,2,2,3,3-D(7) showed a large intramolecular deuterium isotope effect [(k(H)/k(D))(obs)] of 16.0 ± 1.0 for 2-hexanol and 8.9 ± 0.9 for 3-hexanol; and (c) the hydroxylation of the radical clock norcarane led to an estimated radical lifetime of 9.4 ps and an oxygen rebound rate of 1.06 × 10(11) s(-1). These results point to a hydrogen abstraction and oxygen rebound mechanism for alkane hydroxylation. The peroxygenase appeared to lack activity on long-chain alkanes (> C(16)) and highly branched alkanes (e.g. tetramethylpentane), but otherwise exhibited a broad substrate range. It may accordingly have a role in the bioconversion of natural and anthropogenic alkane-containing structures (including alkyl chains of complex biomaterials) in soils, plant litter, and wood.
authors.affiliation: Department of Bio- and Environmental Sciences, International Graduate School of Zittau, Zittau, Germany.
authors.forename: Sebastian
authors.initials: S
authors.lastname: Peter
authors.forename: Matthias
authors.initials: M
authors.lastname: Kinne
authors.forename: Xiaoshi
authors.initials: X
authors.lastname: Wang
authors.forename: René
authors.initials: R
authors.lastname: Ullrich
authors.forename: Gernot
authors.initials: G
authors.lastname: Kayser
authors.forename: John T
authors.initials: JT
authors.lastname: Groves
authors.forename: Martin
authors.initials: M
authors.lastname: Hofrichter
mesh_terms: D055362:Agrocybe / Q000201:enzymology*; D000473:Alkanes / Q000737:chemistry / Q000378:metabolism*; D004795:Enzyme Stability; D006859:Hydrogen / Q000737:chemistry; D006861:Hydrogen Peroxide / Q000737:chemistry / Q000378:metabolism; D006900:Hydroxylation; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D015394:Molecular Structure; D016877:Oxidants / Q000737:chemistry / Q000378:metabolism; D010100:Oxygen / Q000737:chemistry; D012997:Solvents / Q000737:chemistry; D013237:Stereoisomerism
chemical_list: D000473:Alkanes; D016877:Oxidants; D012997:Solvents; D006859:Hydrogen; D006861:Hydrogen Peroxide; D006899:Mixed Function Oxygenases; C022333:peroxygenase; D010100:Oxygen