DOCSLIB.ORG

Oxyfunctionalization of Alkanes, Alkenes and Alkynes by Unspecific Peroxygenase (EC 1.11.2.1) Oxyfunktionalisierung Von Alkanen

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Oxyfunctionalization of Alkanes, Alkenes and Alkynes by Unspecific Peroxygenase (EC 1.11.2.1) Oxyfunktionalisierung Von Alkanen Oxyfunctionalization of alkanes, alkenes and alkynes by unspecific peroxygenase (EC 1.11.2.1) Oxyfunktionalisierung von Alkanen, Alkenen und Alkinen durch die Unspezifische Peroxygenase (EC 1.11.2.1) Vom Institutsrat des Internationalen Hochschulinstitutes Zittau und der Fakultät Mathematik/ Naturwissenschafften der Technischen Universität Dresden genehmigte Dissertation zur Erlangung des akademischen Grades doctor rerum naturalium (Dr. rer. nat.) vorgelegt von Dipl.-Ing. (Umwelttechnik) Sebastian Peter geboren am 04. Juni 1982 in Erlenbach am Main (Deutschland) Gutachter: Herr Prof. Dr. Martin Hofrichter (TU Dresden, IHI Zittau) Herr Prof. Dr. John T. Groves (Princeton University , USA) Frau Prof. Dr. Katrin Scheibner (Hochschule Lausitz) Tag der Verteidigung: 26.04.2013 Oxyfunctionalization of alkanes, alkenes and alkynes by unspecific peroxygenase (EC 1.11.2.1) Approved by the council of International Graduate School of Zittau and the Faculty of Science of the TU Dresden Academic Dissertation Doctor rerum naturalium (Dr. rer. nat.) by Sebastian Peter, Dipl.-Ing. (Environmental Engeneering) born on June 4, 1982 in Erlenbach am Main (Germany) Reviewer: Prof. Dr. Martin Hofrichter (TU Dresden, IHI Zittau) Prof. Dr. John T. Groves (Princeton University , USA) Prof. Dr. Katrin Scheibner (Hochschule Lausitz) Day of defense: April 26, 2013 CONTENTS Contents CONTENTS ............................................................................................................. I LIST OF ABBREVIATIONS .................................................................................. IV ABSTRACT ........................................................................................................... VI 1. INTRODUCTION .............................................................................................. 10 1.1 The importance of enzymes in living organisms ............................................. 10 1.2 Agrocybe aegerita peroxygenase – history and classification over time ........ 10 1.3 Aliphatic hydrocarbons ................................................................................... 21 1.3.1 Alkanes .................................................................................................... 21 1.3.2 Alkenes .................................................................................................... 24 1.3.3. Alkynes ................................................................................................... 25 1.4 Enzymes catalyzing alkane, alkene and alkyne oxidation .............................. 27 1.4.1 Diiron enzymes - methane monooxygenase ............................................ 31 1.4.2 Monoiron enzymes – alkane hydroxylases (alkB) .................................... 33 1.4.3.1 Cytochrome P450 monooxygenases ................................................. 34 1.4.3.2 Heme peroxidases ............................................................................ 38 1.4.4 Biotechnological relevance of hydrocarbon oxygenation and fungal bioremediation .................................................................................................. 40 1.5 Objectives and aims ....................................................................................... 41 2. MATERIALS AND METHODS ......................................................................... 42 2.1 Reagents ........................................................................................................ 42 2.2 Solvent selection and enzyme stability ........................................................... 43 2.2.1 The stability of AaeUPO in organic solvents ............................................ 43 2.2.2 The optimum acetone concentration ........................................................ 43 2.2.3 Inhibition experiment with DMSO ............................................................. 43 2.3 Substrate conversion ...................................................................................... 44 2.3.1 Alkane conversion.................................................................................... 44 2.3.2 Alkene conversion.................................................................................... 44 2.3.3 Alkyne conversion .................................................................................... 45 2.4 Product identification ...................................................................................... 45 2.4.1 Products of alkane conversion ................................................................. 45 2.4.2 Products of alkene and alkyne conversion ............................................... 46 2.4.3 Chiral separation of alcohols, epoxides and alkynols............................... 47 2.5 pH-Optimum of AaeUPO-catalyzed cyclohexane conversion ......................... 47 I 18 2.6 O-labeling experiments ................................................................................ 48 18 2.6.1 Conversion of cyclohexane with H2 O2 ................................................... 48 18 2.6.2 Conversion of 2,3-dimethyl-2-butene with H2 O2 .................................... 48 2.7 Enzyme kinetics ............................................................................................. 49 2.7.1 Cyclohexane conversion .......................................................................... 49 2.7.2 2-Methyl-2-butene conversion ................................................................. 49 2.8 Product quantification ..................................................................................... 49 2.9 Deuterium isotope effect experiments ............................................................ 49 2.10 Radical clock experiments ............................................................................ 50 2.11 Heme N-alkylation by alkenes and alkynes .................................................. 50 2.12 Stopped-flow experiments ............................................................................ 51 2.12.1 Method improvement ............................................................................. 51 2.12.2 AaeUPO compound I – kinetics ............................................................. 51 3. RESULTS ........................................................................................................ 52 3.1 Solvent selection and enzyme stability ........................................................... 52 3.1.1 Stability of AaeUPO in organic solvents .................................................. 52 3.1.2 Inhibition of AaeUPO by DMSO ............................................................... 52 3.2.1 Alkane conversion ................................................................................... 54 3.2.1.1 Linear alkanes ................................................................................... 57 3.2.1.2 Branched alkanes ............................................................................. 57 3.2.1.3 Cyclic alkanes ................................................................................... 57 3.2.1.4 Stereoselectivity of alkane hydroxylation........................................... 60 3.2.2 Alkene conversion ................................................................................... 60 3.2.2.1 Terminal linear alkenes ..................................................................... 61 3.2.2.2 Non-terminal and branched alkenes ................................................. 61 3.2.2.3 Cyclic alkenes ................................................................................... 62 3.2.2.4 Dienes ............................................................................................... 62 3.2.2.5 Stereoselectivity of alkene epoxidation ............................................. 66 3.2.3 Alkyne conversion .................................................................................... 66 3.2.3.1 Linear alkynes ................................................................................... 66 3.2.3.2 Stereoselectivity of alkyne conversion .............................................. 66 3.3 Source of the oxygen incorporated during substrate peroxygenation............. 66 3.4 Kinetic data for AaeUPO-catalyzed reactions ................................................ 69 3.4.1 Cyclohexane hydroxylation ...................................................................... 69 3.4.2 Epoxidation of 2-methyl-2-butene ............................................................ 70 3.5 Deuterium isotope effects ............................................................................... 70 3.6 Evidence for the presence of a radical intermediate....................................... 72 3.7 Enzyme inactivation by heme N-alkylation ..................................................... 73 II CONTENTS 3.8 Stopped-flow experiments .............................................................................. 73 3.8.1 Method improvement ............................................................................... 73 3.8.2 AaeUPO-I – UV/Vis spectrum and properties .......................................... 73 3.8.3 Compound I – kinetic data ....................................................................... 75 4. DISCUSSION ................................................................................................... 75 4.1 AaeUPO stability, inhibition and inactivation .................................................. 78 4.1.1 AaeUPO stability in organic solvents ......................................................
Load more

Recommended publications
  • Dimethyl Sulfoxide MSDS
    He a lt h 1 2 Fire 2 2 0 Re a c t iv it y 0 Pe rs o n a l Pro t e c t io n F Material Safety Data Sheet Dimethyl sulfoxide MSDS Section 1: Chemical Product and Company Identification Product Name: Dimethyl sulfoxide Contact Information: Catalog Codes: SLD3139, SLD1015 Sciencelab.com, Inc. 14025 Smith Rd. CAS#: 67-68-5 Houston, Texas 77396 RTECS: PV6210000 US Sales: 1-800-901-7247 International Sales: 1-281-441-4400 TSCA: TSCA 8(b) inventory: Dimethyl sulfoxide Order Online: ScienceLab.com CI#: Not applicable. CHEMTREC (24HR Emergency Telephone), call: Synonym: Methyl Sulfoxide; DMSO 1-800-424-9300 Chemical Name: Dimethyl Sulfoxide International CHEMTREC, call: 1-703-527-3887 Chemical Formula: (CH3)2SO For non-emergency assistance, call: 1-281-441-4400 Section 2: Composition and Information on Ingredients Composition: Name CAS # % by Weight Dimethyl sulfoxide 67-68-5 100 Toxicological Data on Ingredients: Dimethyl sulfoxide: ORAL (LD50): Acute: 14500 mg/kg [Rat]. 7920 mg/kg [Mouse]. DERMAL (LD50): Acute: 40000 mg/kg [Rat]. Section 3: Hazards Identification Potential Acute Health Effects: Slightly hazardous in case of inhalation (lung irritant). Slightly hazardous in case of skin contact (irritant, permeator), of eye contact (irritant), of ingestion, . Potential Chronic Health Effects: Slightly hazardous in case of skin contact (irritant, sensitizer, permeator), of ingestion. CARCINOGENIC EFFECTS: Not available. MUTAGENIC EFFECTS: Mutagenic for mammalian somatic cells. Mutagenic for bacteria and/or yeast. TERATOGENIC EFFECTS: Not available. DEVELOPMENTAL TOXICITY: Not available. The substance may be toxic to blood, kidneys, liver, mucous membranes, skin, eyes.
    [Show full text]
  • Synthesis of 1-Naphthol by a Natural Peroxygenase Engineered by Directed Evolution
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by UPCommons. Portal del coneixement obert de la UPC Synthesis of 1-Naphthol by a Natural Peroxygenase Engineered by Directed Evolution Patricia Molina-Espeja,[a] Marina Cañellas,[b] Francisco J. Plou,[a] Martin Hofrichter,[c] Fatima Lucas,[b] Victor Guallar[b] and Miguel Alcalde*[a] Abstract: There is an increasing interest in enzymes that catalyze hydroxylation) to the non-natural olefin cyclopropanation by the hydroxylation of naphthalene under mild conditions and with carbene transfer. Indeed, these enzymes can transform minimal requirements. To address this challenge, an extracellular naphthalene into 1-naphthol by either harnessing the peroxide fungal aromatic peroxygenase with mono(per)oxygenase activity shunt pathway or through their natural NAD(P)H dependent was engineered to selectively convert naphthalene into 1-naphthol. activity.[12-15] More recently, directed evolution of toluene ortho- Mutant libraries constructed by random mutagenesis and DNA monooxygenase (TOM) with a whole-cell biocatalytic system recombination were screened for peroxygenase activity on was described.[16-18] However, the poor enzyme stability and the naphthalene while quenching the undesired peroxidative activity on reliance on expensive redox cofactors and reductase domains 1-naphthol (one-electron oxidation). The resulting double mutant have precluded the practical application of these enzymes in (G241D-R257K) of this process was characterized biochemically specific industrial settings. and computationally. The conformational changes produced by directed evolution improved the substrate´s catalytic position. Over a decade ago, the first “true natural” aromatic Powered exclusively by catalytic concentrations of H2O2, this soluble peroxygenase was discovered (EC 1.11.2.1; also referred to as and stable biocatalyst has total turnover numbers of 50,000, with unspecific peroxygenase, UPO).[19] This enzyme was recently high regioselectivity (97%) and reduced peroxidative activity.
    [Show full text]
  • Lpmos) ✉ Riin Kont1, Bastien Bissaro 2,3, Vincent G
    ARTICLE https://doi.org/10.1038/s41467-020-19561-8 OPEN Kinetic insights into the peroxygenase activity of cellulose-active lytic polysaccharide monooxygenases (LPMOs) ✉ Riin Kont1, Bastien Bissaro 2,3, Vincent G. H. Eijsink 2 & Priit Väljamäe 1 Lytic polysaccharide monooxygenases (LPMOs) are widely distributed in Nature, where they catalyze the hydroxylation of glycosidic bonds in polysaccharides. Despite the importance of 1234567890():,; LPMOs in the global carbon cycle and in industrial biomass conversion, the catalytic prop- erties of these monocopper enzymes remain enigmatic. Strikingly, there is a remarkable lack of kinetic data, likely due to a multitude of experimental challenges related to the insoluble nature of LPMO substrates, like cellulose and chitin, and to the occurrence of multiple side reactions. Here, we employed competition between well characterized reference enzymes and LPMOs for the H2O2 co-substrate to kinetically characterize LPMO-catalyzed cellulose oxidation. LPMOs of both bacterial and fungal origin showed high peroxygenase efficiencies, 5 6 −1 −1 with kcat/KmH2O2 values in the order of 10 –10 M s . Besides providing crucial insight into the cellulolytic peroxygenase reaction, these results show that LPMOs belonging to multiple families and active on multiple substrates are true peroxygenases. 1 Institute of Molecular and Cell Biology, University of Tartu, Tartu, Estonia. 2 Faculty of Chemistry, Biotechnology and Food Science, NMBU—Norwegian University of Life Sciences, Ås, Norway. 3 INRAE, Aix Marseille University,
    [Show full text]
  • A Flexible All-Atom Model of Dimethyl Sulfoxide for Molecular Dynamics Simulations
    1074 J. Phys. Chem. A 2002, 106, 1074-1080 A Flexible All-Atom Model of Dimethyl Sulfoxide for Molecular Dynamics Simulations Matthew L. Strader and Scott E. Feller* Department of Chemistry, Wabash College, CrawfordsVille, Indiana 47933 ReceiVed: October 1, 2001 An all-atom, flexible dimethyl sulfoxide model has been created for molecular dynamics simulations. The new model was tested against experiment for an array of thermodynamic, structural, and dynamic properties. Interactions with water were compared with previous simulations and experimental studies, and the unusual changes exhibited by dimethyl sulfoxide/water mixtures, such as the enhanced structure of the solution, were reproduced by the new model. Particular attention was given to the design of the electrostatic component of the force field and to providing compatibility with the CHARMM parameter sets for biomolecules. Introduction mixtures,14,17,18 and its interaction with a phospholipid bi- Theoretical methods for the investigation of biological layer.19,20 Simulations published to date have utilized a rigid, molecules have become commonplace in biochemistry and united-atom model where each methyl group is represented by biophysics.1 These approaches provide detailed atomic models a single atomic center and all bond lengths and angles are fixed of biological systems, from which structure-function relation- at their equilibrium values. Another feature common to most ships can be elucidated. Methods based on empirical force fields, of these models is the use of the same charge distribution, such as molecular dynamics (MD) simulations, allow for namely that obtained by Rao and Singh from a Hartree-Fock calculations on relatively large systems, e.g., complete biomol- 6-31G* ab initio quantum mechanical calculation.21 ecules or assemblies of biomolecules in their aqueous environ- Recent advances in computer hardware now allow a flexible, ment.
    [Show full text]
  • © Central University of Technology, Free State DECLARATION
    © Central University of Technology, Free State DECLARATION DECLARATION I, MOHAMMAD PARVEZ (INDIAN PASSPORT NUMBER ), hereby certify that the dissertation submitted by me for the degree DOCTOR OF HEALTH SCIENCES IN BIOMEDICAL TECHNOLOGY, is my own independent work; and complies with the Code of Academic Integrity, as well as other relevant policies, procedures, rules and regulations of the Central University of Technology (Free State). I hereby declare, that this research project has not been previously submitted before to any university or faculty for the attainment of any qualification. I further waive copyright of the dissertation in favour of the Central University of Technology (Free State). I also state that expression vector modified/generated in this study is in collaboration with my colleagues, Mr HANS DENIS BAMAL (student number: ) and Ms IPELENG KOPANO ROSINAH KGOSIEMANG (student number: ). MOHAMMAD PARVEZ DATE II | P a g e © Central University of Technology, Free State © Central University of Technology, Free State ACKNOWLEDGEMENT ACKNOWLEDGEMENTS This thesis would have remained a dream, had it not been for my supervisor, Prof Khajamohiddin Syed, who continuously supported me during my doctoral study. I would like to extend my gratitude to him, for his patience, motivation, enthusiasm, and immense knowledge in P450 research. I would also like to thank him for giving me the opportunity to attend national and international scientific conferences. He has shown great faith and trust in me throughout my studies and without such attributes I may not be where I am now. I would also like to thank Prof Samson Sitheni Mashele for his support, and always listening to our concerns.
    [Show full text]
  • Selective Synthesis of the Human Drug Metabolite 5′-Hydroxypropranolol by an Evolved Self-Sufficient Peroxygenase
    This is an open access article published under an ACS AuthorChoice License, which permits copying and redistribution of the article or any adaptations for non-commercial purposes. Research Article Cite This: ACS Catal. 2018, 8, 4789−4799 pubs.acs.org/acscatalysis Selective Synthesis of the Human Drug Metabolite 5′- Hydroxypropranolol by an Evolved Self-Sufficient Peroxygenase † ‡ § § Patricia Gomez de Santos, Marina Cañellas, Florian Tieves, Sabry H. H. Younes, † ∥ § ‡ † Patricia Molina-Espeja, Martin Hofrichter, Frank Hollmann, Victor Guallar, and Miguel Alcalde*, † Department of Biocatalysis, Institute of Catalysis, CSIC, 28049 Madrid, Spain ‡ Joint BSC-CRG-IRB Research Program in Computational Biology, Barcelona Supercomputing Center, 08034 Barcelona, Spain § Department of Biotechnology, Delft University of Technology, van der Massweg 9, 2629HZ Delft, The Netherlands ∥ Department of Bio- and Environmental Sciences, TU Dresden, International Institute Zittau, Mark 23, 02763 Zittau, Germany *S Supporting Information ABSTRACT: Propranolol is a widely used beta-blocker that is metabolized by human liver P450 monooxygenases into equipotent hydroxylated human drug metabolites (HDMs). It is paramount for the pharmaceutical industry to evaluate the toxicity and activity of these metabolites, but unfortunately, their synthesis has hitherto involved the use of severe conditions, with poor reaction yields and unwanted by- products. Unspecific peroxygenases (UPOs) catalyze the selective oxyfunctionalization of C−H bonds, and they are of particular interest in synthetic organic chemistry. Here, we describe the engineering of UPO from Agrocybe aegerita for the efficient synthesis of 5′-hydroxypropranolol (5′-OHP). We employed a structure-guided evolution approach combined with computational analysis, with the aim of avoiding unwanted phenoxyl radical coupling without having to dope the reaction with radical scavengers.
    [Show full text]
  • Dimethyl Sulfoxide Oxidation of Primary Alcohols
    Western Michigan University ScholarWorks at WMU Master's Theses Graduate College 8-1966 Dimethyl Sulfoxide Oxidation of Primary Alcohols Carmen Vargas Zenarosa Follow this and additional works at: https://scholarworks.wmich.edu/masters_theses Part of the Chemistry Commons Recommended Citation Zenarosa, Carmen Vargas, "Dimethyl Sulfoxide Oxidation of Primary Alcohols" (1966). Master's Theses. 4374. https://scholarworks.wmich.edu/masters_theses/4374 This Masters Thesis-Open Access is brought to you for free and open access by the Graduate College at ScholarWorks at WMU. It has been accepted for inclusion in Master's Theses by an authorized administrator of ScholarWorks at WMU. For more information, please contact [email protected]. DIMETHYL SULFOXIDE OXIDATION OF PRIMARY ALCOHOLS by Carmen Vargas Zenarosa A thesis presented to the Faculty of the School of Graduate Studies in partial fulfillment of the Degree of Master of Arts Western Michigan University Kalamazoo, Michigan August, 1966 ACKNOWLEDGMENTS The author wishes to express her appreciation to the members of her committee, Dr, Don C. Iffland and Dr. Donald C, Berndt, for their helpful suggestions and most especially to Dr, Robert E, Harmon for his patience, understanding, and generous amount of time given to insure the completion of this work. Appreciation is also expressed for the assistance given by her. colleagues. The author acknowledges the assistance given by the National Institutes 0f Health for this research project. Carmen Vargas Zenarosa ii TABLE OF CONTENTS Page
    [Show full text]
  • Evaluation of Yarrowia Lipolytica As a Host For
    EVALUATION OF YARROWIA LIPOLYTICA AS A HOST FOR CYTOCHROME P450 MONOOXYGENASE EXPRESSION By GEORGE OGELLO OBIERO B.Sc. (U.O.N); M.Sc. (U.B) Submitted in fulfilment of the requirements for the degree PHILOSOPHIAE DOCTOR In the Faculty of Natural and Agricultural Sciences, Department of Microbial, Biochemical and Food Biotechnology at the University of the Free State, Bloemfontein, South Africa June, 2006 Promoter: Prof. M.S. Smit "It's not that I'm so smart, it's just that I stay with problems longer." Albert Einstein (1879-1955) I This thesis is dedicated to my late mother, Mrs. Beldine Obiero and my dad, Mr. Gabriel Obiero whose unwavering support and encouragement has been my pillar of strength. II ACKNOWLEDGEMENTS I would like to express my sincere gratitude toward the following people: Prof. M.S. Smit for her invaluable guidance, patience and constructive criticism during the course of the study. Thanks for the patience and determination throughout this project. Mr. P.J. Botes for his technical assistance with the chemical analyses. Dr. E. setati for her valuable and motivating discussions. All my colleagues in Biotransformation Research Group. Fellow students and staff in the Department of Microbial, Biochemical and Food Biotechnology, University of Free State My family and friends for being there for me throughout the study and for their much needed words of encouragement and support. National Research Foundation (NRF) for the financial support of this project. The Heavenly Father without whom this work would not have been successful. III TABLE OF CONTENTS CHAPTER 1: INTRODUCTION 1. Introduction............................................................................................................ 1 1.1.
    [Show full text]
  • Structural Insights Into the Substrate Promiscuity of a Laboratory-Evolved Peroxygenase
    Articles Cite This: ACS Chem. Biol. 2018, 13, 3259−3268 pubs.acs.org/acschemicalbiology Structural Insights into the Substrate Promiscuity of a Laboratory-Evolved Peroxygenase † ∥ ‡ ∥ ‡ Mercedes Ramirez-Escudero, , Patricia Molina-Espeja, , Patricia Gomez de Santos, § † ‡ Martin Hofrichter, Julia Sanz-Aparicio,*, and Miguel Alcalde*, † Department of Crystallography & Structural Biology, Institute of Physical Chemistry “Rocasolano”, CSIC, 28006 Madrid, Spain ‡ Department of Biocatalysis, Institute of Catalysis, CSIC, 28049 Madrid, Spain § Department of Bio- and Environmental Sciences, TU Dresden, International Institute Zittau, Mark 23, 02763 Zittau, Germany *S Supporting Information ABSTRACT: Because of their minimal requirements, sub- strate promiscuity and product selectivity, fungal peroxyge- nases are now considered to be the jewel in the crown of C−H oxyfunctionalization biocatalysts. In this work, the crystal struc- ture of the first laboratory-evolved peroxygenase expressed by yeast was determined at a resolution of 1.5 Å. Notable differ- ences were detected between the evolved and native peroxy- genase from Agrocybe aegerita, including the presence of a full N-terminus and a broader heme access channel due to the mutations that accumulated through directed evolution. Fur- ther mutagenesis and soaking experiments with a palette of peroxygenative and peroxidative substrates suggested dynamic trafficking through the heme channel as the main driving force for the exceptional substrate promiscuity of peroxygenase. Accordingly, this study
    [Show full text]
  • Europe PMC Funders Group Author Manuscript Nat Catal
    Europe PMC Funders Group Author Manuscript Nat Catal. Author manuscript; available in PMC 2018 May 20. Published in final edited form as: Nat Catal. 2018 January ; 1(1): 55–62. doi:10.1038/s41929-017-0001-5. Europe PMC Funders Author Manuscripts Selective aerobic oxidation reactions using a combination of photocatalytic water oxidation and enzymatic oxyfunctionalisations Wuyuan Zhang[a], Elena Fernández-Fueyo[a], Yan Ni[a], Morten van Schie[a], Jenö Gacs[a], Rokus Renirie[b], Ron Wever[b], Francesco G. Mutti[b], Dörte Rother[c], Miguel Alcalde[d], and Frank Hollmann[a],* [a]Department of Biotechnology, Delft University of Technology, Van der Maasweg 9, 2629HZ Delft, The Netherlands [b]Van’t Hoff Institute for Molecular Sciences (HIMS), Faculty of Science, University of Amsterdam, Science Park 904, 1098 XH Amsterdam, The Netherlands [c]Institute of Bio- and Geosciences, IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, 52425 Jülich, Germany [d]Department of Biocatalysis, Institute of Catalysis, CSIC, 28049 Madrid, Spain Abstract Peroxygenases offer attractive means to address challenges in selective oxyfunctionalisation chemistry. Despite their attractiveness, the application of peroxygenases in synthetic chemistry remains challenging due to their facile inactivation by the stoichiometric oxidant (H2O2). Often atom inefficient peroxide generation systems are required, which show little potential for large Europe PMC Funders Author Manuscripts scale implementation. Here we show that visible light-driven, catalytic water oxidation can be used for in situ generation of H2O2 from water, rendering the peroxygenase catalytically active. In this way the stereoselective oxyfunctionalisation of hydrocarbons can be achieved by simply using the catalytic system, water and visible light.
    [Show full text]
  • Targeting of Insect Epicuticular Lipids by the Entomopathogenic Fungus Beauveria Bassiana: Hydrocarbon Oxidation Within the Context of a Host-Pathogen Interaction
    ORIGINAL RESEARCH ARTICLE published: 15 February 2013 doi: 10.3389/fmicb.2013.00024 Targeting of insect epicuticular lipids by the entomopathogenic fungus Beauveria bassiana: hydrocarbon oxidation within the context of a host-pathogen interaction Nicolás Pedrini 1, Almudena Ortiz-Urquiza 2, Carla Huarte-Bonnet 1, Shizhu Zhang 2,3 and Nemat O. Keyhani 2* 1 Facultad de Ciencias Médicas, Instituto de Investigaciones Bioquímicas de La Plata (CCT La Plata CONICET-UNLP), La Plata, Argentina 2 Department of Microbiology and Cell Science, University of Florida, Gainesville, FL, USA 3 Jiangsu Key Laboratory for Microbes and Functional Genomics, Jiangsu Engineering and Technology Research Center for Microbiology, College of Life Sciences, Nanjing Normal University, Nanjing, China Edited by: Broad host range entomopathogenic fungi such as Beauveria bassiana attack insect hosts Rachel N. Austin, Bates College, via attachment to cuticular substrata and the production of enzymes for the degradation USA and penetration of insect cuticle. The outermost epicuticular layer consists of a complex Reviewed by: mixture of non-polar lipids including hydrocarbons, fatty acids, and wax esters. Long chain Rachel N. Austin, Bates College, USA hydrocarbons are major components of the outer waxy layer of diverse insect species, Alan A. DiSpirito, Ohio State where they serve to protect against desiccation and microbial parasites, and as recognition University, USA molecules or as a platform for semiochemicals. Insect pathogenic fungi have evolved *Correspondence: mechanisms for overcoming this barrier, likely with sets of lipid degrading enzymes with Nemat O. Keyhani, Department of overlapping substrate specificities. Alkanes and fatty acids are substrates for a specific Microbiology and Cell Science, University of Florida, Gainesville, subset of fungal cytochrome P450 monooxygenases involved in insect hydrocarbon FL 32611, USA.
    [Show full text]
  • A Comparative Genomic Investigation of Niche Adaptation in Fungi”
    The Pennsylvania State University The Graduate School Huck Institute of the Life Sciences A COMPARATIVE GENOMIC INVESTIGATION OF NICHE ADAPTATION IN FUNGI A Dissertation in Integrative Biosciences by Venkatesh Moktali © 2013 Venkatesh Moktali Submitted in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy August 2013 ! The dissertation of Venkatesh Moktali was reviewed and approved* by the following: Seogchan Kang Professor of Plant Pathology and Environmental Microbiology Dissertation Advisor Chair of Committee David M. Geiser Professor of Plant Pathology and Environmental Microbiology Kateryna Makova Professor of Biology Anton Nekrutenko Associate Professor of Biochemistry and Molecular Biology Yu Zhang Associate Professor of Statistics Peter Hudson Department Head, Huck Institute of the Life Sciences *Signatures are on file in the Graduate School ! ! Abstract The Kingdom Fungi has a diverse array of members adapted to very disparate and the most hostile surroundings on earth: such as living plant and/or animal tissues, soil, aquatic environments, other microorganisms, dead animals, and exudates of plants, animals and even nuclear reactors. The ability of fungi to survive in these various niches is supported by the presence of key enzymes/proteins that can metabolize extraneous harmful factors. Characterization of the evolution of these key proteins gives us a glimpse at the molecular mechanisms underpinning adaptations in these organisms. Cytochrome P450 proteins (CYPs) are among the most diversified protein families, they are involved in a number of processed that are critical to fungi. I evaluated the evolution of Cytochrome P450 proteins (CYPs) in order to understand niche adaptation in fungi. Towards this goal, a previously developed database the fungal cytochrome P450 database (FCPD) was improved and several features were added in order to allow for systematic comparative genomic and phylogenomic analysis of CYPs from numerous fungal genomes.
    [Show full text]