Steroid hydroxylation by basidiomycete peroxygenases: A combined experimental and computational Study

Autor: Victor Guallar, Marina Cañellas, José C. del Río, Katrin Scheibner, Lisbeth Kalum, Glenn Gröbe, René Ullrich, Ferran Sancho, Henrik Lund, Martin Hofrichter, Ángel T. Martínez, Ana Gutiérrez, Fátima Lucas, Esteban D. Babot
Rok vydání: 2015
Předmět:
Conjugated double bonds
medicine.medical_treatment
Computational studies
Recombinant enzymes
01 natural sciences
Applied Microbiology and Biotechnology
Mixed Function Oxygenases
Hydroxylation
chemistry.chemical_compound
Coprinopsis cinerea
Side chain
Organic chemistry
Steroidal compounds
0303 health sciences
Gas chromatography
Ecology
biology
Chains
Stereoisomerism
Environmentally friendly condition
Ketones
Ligand (biochemistry)
Lipids
Marasmius
Enzymes
Steroids
Biotechnology
Chromatography
Gas
Stereochemistry
Enzyme catalysis
Steroid
03 medical and health sciences
Organic compounds
medicine
Computer Simulation
Enzymology and Protein Engineering
030304 developmental biology
Mass spectrometry
010405 organic chemistry
Agrocybe
Fungi
Hydrogen Peroxide
Oxyfunctionalization
biology.organism_classification
0104 chemical sciences
chemistry
Protein energy landscape
Agaricales
Food Science
Zdroj: Digital.CSIC. Repositorio Institucional del CSIC
instname
Popis: 13 páginas.-- 7 figuras.-- 4 tablas.-- 40 referencias.-- Supplemental material for this article may be found at http://dx.doi.org/10.1128/AEM.00660-15.
The goal of this study is the selective oxyfunctionalization of steroids under mild and environmentally friendly conditions using fungal enzymes. With this purpose, peroxygenases from three basidiomycete species were tested for the hydroxylation of a variety of steroidal compounds, using H2O2 as the only cosubstrate. Two of them are wild-type enzymes from Agrocybe aegerita and Marasmius rotula, and the third one is a recombinant enzyme from Coprinopsis cinerea. The enzymatic reactions on free and esterified sterols, steroid hydrocarbons, and ketones were monitored by gas chromatography, and the products were identified by mass spectrometry. Hydroxylation at the side chain over the steroidal rings was preferred, with the 25-hydroxyderivativespredominating. Interestingly, antiviral and other biological activities of 25-hydroxycholesterol have been reported recently (M. Blanc et al., Immunity 38:106-118, 2013, http://dx.doi.org/10.1016/j.immuni.2012.11.004). However, hydroxylation in the ring moiety and terminal hydroxylation at the side chain also was observed in some steroids, the former favored by the absence of oxygenated groups at C-3 and by the presence of conjugated double bonds in the rings. To understand the yield and selectivity differences between the different steroids, a computational study was performed using Protein Energy Landscape Exploration (PELE) software for dynamic ligand diffusion. These simulations showed that the active-site geometry and hydrophobicity favors the entrance of the steroid side chain, while the entrance of the ring is energetically penalized. Also, a direct correlation between the conversion rate and the side chain entrance ratio could be established that explains the various reaction yields observed.
This study was supported by the INDOX (KBBE-2013-7-613549), PEROXICATS (KBBE-2010-4-265397), and PELE (ERC-2009-Adg 25027) EU projects. We have no conflicts of interest to declare.
Databáze: OpenAIRE
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