Genome-to-function characterization of novel fungal P450 monooxygenases oxidizing polycyclic aromatic hydrocarbons (PAHs).

TitleGenome-to-function characterization of novel fungal P450 monooxygenases oxidizing polycyclic aromatic hydrocarbons (PAHs).
Publication TypeJournal Article
Year of Publication2010
AuthorsSyed, K, Doddapaneni, H, Subramanian, V, Lam, YWai, Yadav, JS
JournalBiochem Biophys Res Commun
Volume399
Issue4
Pagination492-7
Date Published2010 Sep 03
ISSN1090-2104
KeywordsBiodegradation, Environmental, Catalysis, Cytochrome P-450 Enzyme System, Genome, Fungal, Genome-Wide Association Study, Mixed Function Oxygenases, Oxidation-Reduction, Phanerochaete, Pichia, Polycyclic Aromatic Hydrocarbons, Recombinant Proteins, Substrate Specificity
Abstract

Fungi, particularly the white rot basidiomycetes, have an extraordinary capability to degrade and/or mineralize (to CO(2)) the recalcitrant fused-ring high molecular weight (4 aromatic-rings) polycyclic aromatic hydrocarbons (HMW PAHs). Despite over 30years of research demonstrating involvement of P450 monooxygenation reactions in fungal metabolism of HMW PAHs, specific P450 monooxygenases responsible for oxidation of these compounds are not yet known. Here we report the first comprehensive identification and functional characterization of P450 monooxygenases capable of oxidizing different ring-size PAHs in the model white rot fungus Phanerochaete chrysosporium using a successful genome-to-function strategy. In a genome-wide P450 microarray screen, we identified six PAH-responsive P450 genes (Pc-pah1-Pc-pah6) inducible by PAHs of varying ring size, namely naphthalene, phenanthrene, pyrene, and benzo(a)pyrene (BaP). Using a co-expression strategy, cDNAs of the six Pc-Pah P450s were cloned and expressed in Pichia pastoris in conjunction with the homologous P450 oxidoreductase (Pc-POR). Each of the six recombinant P450 monooxygenases showed PAH-oxidizing activity albeit with varying substrate specificity towards PAHs (3-5 rings). All six P450s oxidized pyrene (4-ring) into two monohydroxylated products. Pc-Pah1 and Pc-Pah3 oxidized BaP (5-ring) to 3-hydroxyBaP whereas Pc-Pah4 and Pc-Pah6 oxidized phenanthrene (3-ring) to 3-, 4-, and 9-phenanthrol. These PAH-oxidizing P450s (493-547 aa) are structurally diverse and novel considering their low overall homology (12-23%) to mammalian counterparts. To our knowledge, this is the first report on specific fungal P450 monooxygenases with catalytic activity toward environmentally persistent and highly toxic HMW PAHs.

DOI10.1016/j.bbrc.2010.07.094
Alternate JournalBiochem. Biophys. Res. Commun.
PubMed ID20674550
PubMed Central IDPMC2943217
Grant ListR01 ES015543-01A1 / ES / NIEHS NIH HHS / United States
R01 ES010210 / ES / NIEHS NIH HHS / United States
R01ES015543 / ES / NIEHS NIH HHS / United States
R01 ES015543 / ES / NIEHS NIH HHS / United States
R01 ES010210-01A1 / ES / NIEHS NIH HHS / United States
R01ES10210 / ES / NIEHS NIH HHS / United States