@misc{KiebistSchmidtkeZimmermannetal., author = {Kiebist, Jan and Schmidtke, Kai-Uwe and Zimmermann, J{\"o}rg and Kellner, Harald and Jehmlich, Nico and Ullrich, Ren{\´e} and Z{\"a}nder, Daniel and Hofrichter, Martin and Scheibner, Katrin}, title = {A peroxygenase from Chaetomium globosum catalyzes the selective oxygenation of testosterone}, series = {ChemBioChem}, volume = {18}, journal = {ChemBioChem}, number = {6}, issn = {1439-7633}, doi = {10.1002/cbic.201600677}, pages = {563 -- 569}, abstract = {Unspecific peroxygenases (UPO, EC 1.11.2.1) secreted by fungi open an efficient way to selectively oxyfunctionalize diverse organic substrates including less activated hydrocarbons by transferring peroxide-borne oxygen. Herein, we investigated a cell-free approach to incorporate epoxy and hydroxyl functionalities directly into the bulky molecule of testosterone by a novel unspecific peroxygenase that was produced by the ascomycetous fungus Chaetomium globosum in a complex medium rich in carbon and nitrogen. Purification by fast protein liquid chromatography revealed two enzyme fractions with the same molecular mass of 36 kDa and specific activities of 4.4 to 12 U mg-1. Whereas well-known UPOs of Agrocybe aegerita (AaeUPO) and Marasmius rotula (MroUPO) failed to convert testosterone in a comparative study, the UPO of C. globosum (CglUPO) accepted testosterone as substrate and converted it with up to 7,000 total turnovers (TTN) into two oxygenated products: the 4,5-epoxide of testosterone in β-configuration and 16α-hydroxytestosterone. The reaction was performed at 100-mg scale resulting in the formation of about 90 \% of the epoxide and 10 \% of the hydroxylation product, which both could be isolated with purities above 96 \%. Thus, CglUPO may be a promising biocatalyst for the oxyfunctionalization of bulky steroids and provide a useful tool for the synthesis of pharmaceutically relevant steroidal molecules.}, language = {en} } @misc{KarichUllrichScheibneretal., author = {Karich, Alexander and Ullrich, Ren{\´e} and Scheibner, Katrin and Hofrichter, Martin}, title = {Fungal unspecific peroxygenases oxidize the majority of organic EPA priority pollutants}, series = {Frontiers in Microbiology}, volume = {8}, journal = {Frontiers in Microbiology}, issn = {1664-302X}, doi = {10.3389/fmicb.2017.01463}, pages = {15}, abstract = {Unspecific peroxygenases (UPOs) are secreted fungal enzymes with promiscuity for oxygen transfer and oxidation reactions. Functionally, they represent hybrids of P450 monooxygenases and heme peroxidases; phylogenetically they belong to the family of heme-thiolate peroxidases. Two UPOs from the basidiomycetous fungi Agrocybe aegerita (AaeUPO) and Marasmius rotula (MroUPO) converted 35 out of 40 compounds listed as EPA priority pollutants, including chlorinated benzenes and their derivatives, halogenated biphenyl ethers, nitroaromatic compounds, polycyclic aromatic hydrocarbons (PAHs) and phthalic acid derivatives. These oxygenations and oxidations resulted in diverse products and-if at all-were limited for three reasons: (i) steric hindrance caused by multiple substitutions or bulkiness of the compound as such (e.g., hexachlorobenzene or large PAHs), (ii) strong inactivation of aromatic rings (e.g., nitrobenzene), and (iii) low water solubility (e.g., complex arenes). The general outcome of our study is that UPOs can be considered as extracellular counterparts of intracellular monooxygenases, both with respect to catalyzed reactions and catalytic versatility. Therefore, they should be taken into consideration as a relevant biocatalytic detoxification and biodegradation tool used by fungi when confronted with toxins, xenobiotics and pollutants in their natural environments.}, language = {en} } @misc{OlmedoRioKiebistetal., author = {Olmedo, Andr{\´e}s and R{\´i}o, Jos{\´e} C. del and Kiebist, Jan and Ullrich, Ren{\´e} and Hofrichter, Martin and Scheibner, Katrin and Mart{\´i}nez, Angel T. and Guti{\´e}rrez, Ana}, title = {Fatty Acid Chain Shortening by a Fungal Peroxygenase}, series = {Chemistry A European Journal}, volume = {23}, journal = {Chemistry A European Journal}, issn = {1521-3765}, doi = {10.1002/chem.201704773}, pages = {16989 -- 67}, abstract = {A recently discovered peroxygenase from the fungus Marasmius rotula (MroUPO) is able to catalyze the progressive one-carbon shortening of medium and longchain mono- and dicarboxylic acids by itself alone, in the presence of H₂O₂. The mechanism, analyzed using H₂O, starts with an a-oxidation catalyzed by MroUPO generat- ing an α-hydroxy acid, which is further oxidized by the enzyme to a reactive α-keto intermediate whose decarboxylation yields the one-carbon shorter fatty acid. Compared with the previously characterized peroxygenase of Agrocybe aegerita, a wider heme access channel, enabling fatty acid positioning with the carboxylic end near the heme cofactor (as seen in one of the crystal structures available) could be at the origin of the unique ability of MroUPO shortening carboxylic acid chains.}, language = {en} } @misc{MartinezRuizDuenasCamareroetal., author = {Martinez, Angel T. and Ruiz-Duenas, Francisco J. and Camarero, Susana and Serrano, Ana and Linde, Dolores and Lund, Henrik and Vind, Jesper and Tovborg, Morton and Herold-Majumdar, Owik M. and Hofrichter, Martin and Liers, Christiane and Ullrich, Ren{\´e} and Scheibner, Katrin and Sannia, Giovanni and Piscitelli, Alessandra and Sener, Mehmet E. and Kilic, Sibel and Berkel, Willem J. H. van and Guallar, V{\´i}ctor and Lucas, Maria F{\´a}tima and Zuhse, Ralf and Ludwig, Roland and Hollmann, Frank and Fern{\´a}ndez-Fueyo, Elena and Record, Eric and Faulds, Craig B. and Tortajada, Marta and Winckelmann, Ib and Rasmussen, Jo-Anne and Gelo-Pujic, Mirjana and Guti{\´e}rrez, Ana and Rio, Jos{\´e} C. del and Rencoret, Jorge and Alcalde, Miguel}, title = {Oxidoreductases on their way to industrial biotransformations}, series = {Biotechnology Advances}, volume = {35}, journal = {Biotechnology Advances}, number = {6}, issn = {1873-1899}, doi = {10.1016/j.biotechadv.2017.06.003}, pages = {815 -- 831}, abstract = {Fungi produce heme-containing peroxidases and peroxygenases, flavin-containing oxidases and dehydrogenases, and different copper-containing oxidoreductases involved in the biodegradation of lignin and other recalcitrant compounds. Heme peroxidases comprise the classical ligninolytic peroxidases and the new dye-decolorizing peroxidases, while heme peroxygenases belong to a still largely unexplored superfamily of heme-thiolate proteins. Nevertheless, basidiomycete unspecific peroxygenases have the highest biotechnological interest due to their ability to catalyze a variety of regio- and stereo-selective monooxygenation reactions with H2O2 as the source of oxygen and final electron acceptor. Flavo-oxidases are involved in both lignin and cellulose decay generating H2O2 that activates peroxidases and generates hydroxyl radical. The group of copper oxidoreductases also includes other H2O2 generating enzymes - copper-radical oxidases - together with classical laccases that are the oxidoreductases with the largest number of reported applications to date. However, the recently described lytic polysaccharide monooxygenases have attracted the highest attention among copper oxidoreductases, since they are capable of oxidatively breaking down crystalline cellulose, the disintegration of which is still a major bottleneck in lignocellulose biorefineries, along with lignin degradation. Interestingly, some flavin-containing dehydrogenases also play a key role in cellulose breakdown by directly/indirectly "fueling" electrons for polysaccharide monooxygenase activation. Many of the above oxidoreductases have been engineered, combining rational and computational design with directed evolution, to attain the selectivity, catalytic efficiency and stability properties required for their industrial utilization. Indeed, using ad hoc software and current computational capabilities, it is now possible to predict substrate access to the active site in biophysical simulations, and electron transfer efficiency in biochemical simulations, reducing in orders of magnitude the time of experimental work in oxidoreductase screening and engineering. What has been set out above is illustrated by a series of remarkable oxyfunctionalization and oxidation reactions developed in the frame of an intersectorial and multidisciplinary European RTD project. The optimized reactions include enzymatic synthesis of 1-naphthol, 25-hydroxyvitamin D3, drug metabolites, furandicarboxylic acid, indigo and other dyes, and conductive polyaniline, terminal oxygenation of alkanes, biomass delignification and lignin oxidation, among others. These successful case stories demonstrate the unexploited potential of oxidoreductases in medium and large-scale biotransformations.}, language = {en} }