CYP63A2, a Catalytically Versatile Fungal P450 Monooxygenase Capable of Oxidizing Higher-Molecular-Weight Polycyclic Aromatic Hydrocarbons, Alkylphenols, and Alkanes

Abstract

c Cytochrome P450 monooxygenases (P450s) are known to oxidize hydrocarbons, albeit with limited substrate specificity across classes of these compounds. Here we report a P450 monooxygenase (CYP63A2) from the model ligninolytic white rot fungus Phanerochaete chrysosporium that was found to possess a broad oxidizing capability toward structurally diverse hydrocarbons belonging to mutagenic/carcinogenic fused-ring higher-molecular-weight polycyclic aromatic hydrocarbons (HMW-PAHs), endocrine-disrupting long-chain alkylphenols (APs), and crude oil aliphatic hydrocarbon n-alkanes. A homology-based threedimensional (3D) model revealed the presence of an extraordinarily large active-site cavity in CYP63A2 compared to the mammalian PAH-oxidizing (CYP3A4, CYP1A2, and CYP1B1) and bacterial aliphatic-hydrocarbon-oxidizing (CYP101D and CYP102A1) P450s. This structural feature in conjunction with ligand docking simulations suggested potential versatility of the enzyme. Experimental characterization using recombinantly expressed CYP63A2 revealed its ability to oxidize HMW-PAHs of various ring sizes, including 4 rings (pyrene and fluoranthene), 5 rings [benzo(a)pyrene], and 6 rings [benzo(ghi)perylene], with the highest enzymatic activity being toward the 5-ring PAH followed by the 4-ring and 6-ring PAHs, in that order. Recombinant CYP63A2 activity yielded monohydroxylated PAH metabolites. The enzyme was found to also act as an alkane -hydroxylase that oxidized n-alkanes with various chain lengths (C 9 to C 12 and C 15 to C 19 ), as well as alkyl side chains (C 3 to C 9 ) in alkylphenols (APs). CYP63A2 showed preferential oxidation of long-chain APs and alkanes. To our knowledge, this is the first P450 identified from any of the biological kingdoms that possesses such broad substrate specificity toward structurally diverse xenobiotics (PAHs, APs, and alkanes), making it a potent enzyme biocatalyst candidate to handle mixed pollution (e.g., crude oil spills).

Enzymatic Catalysis of Hydrocarbons Oxidation “in vitro” (Review)

“…It should be noted that one enzyme was specified [42], Cytochrome CYP63A2, which was estimated as the first P450 identified from any of the biological kingdoms that possesses broad substrate specificity toward structurally diverse xenobioticshigher-molecular-weight polycyclic aromatic hydrocarbons, long-chain alkylphenols, and crude oil aliphatic hydrocarbon n-alkanes. It makes this enzyme an effective handler of mixed hydrocarbon pollutions, e.g.…”

“…This structural feature in conjunction with ligand docking simulations suggested potential versatility of the enzyme. Experimental characterization using recombinantly expressed CYP63A2 revealed its ability to oxidize HMW-PAHs of various ring sizes, including 4 rings (pyrene and fluoranthene), 5 rings [benzo(α)pyrene], and 6 rings [benzo(ghi) perylene], with the highest enzymatic activity being toward the 5-ring PAH followed by the 4-ring and 6-ring PAHs, in that order [42]. Recombinant CYP63A2 activity yielded monohydroxylated PAHs metabolites.…”

“…Cytochrome P450 monooxygenase (CYP63A2) from the model ligninolytic white rot fungus Phanerochaete chrysosporium, that was found to possess a broad oxidizing capability toward structurally diverse hydrocarbons belonging to mutagenic/carcinogenic fusedring higher-molecular-weight polycyclic aromatic hydrocarbons (HMW-PAHs), endocrine-disrupting longchain alkylphenols (APs), and crude oil aliphatic hydrocarbon n-alkanes was reported [42]. A homologybased three-dimensional (3D) model revealed the presence of an extraordinarily large active-site cavity in CYP63A2 compared to the mammalian PAH-oxidizing (CYP3A4, CYP1A2, and CYP1B1) and bacterial aliphatic-hydrocarbon-oxidizing (CYP101D and CYP102A1) P450s.…”

“…The authors postulated that this is the first P450 identified from any of the biological kingdoms that possesses such broad substrate specificity toward structurally diverse xenobiotics (PAHs, APs, and alkanes), making it a potent enzyme biocatalyst candidate to handle mixed pollution, e.g. crude oil spills [42].…”

Szénhidrogén bontó Rhodococcus erythropolis törzsek funkcionális genomikai analízise és biotechnológiai alkalmazása

“…A CYP52-es családba tartozó monooxigenázok katalizálják a zsírsavak ω-hidroxilációját Candida fajokban. Az ugyancsak gomba eredetű CYP63A2 típusú monooxigenáz széles szubsztrát spektrummal rendelkezik, így egyaránt képes az alkánok, PAH-ok és alkil-fenolok oxidációjára [51].…”