Expression of the Caldariomyces fumagoChloroperoxidase in Aspergillus niger and Characterization of the Recombinant Enzyme*

The Caldariomyces fumagochloroperoxidase was successfully expressed in Aspergillus niger. The recombinant enzyme was produced in the culture medium as an active protein and could be purified by a three-step purification procedure. The catalytic behavior of recombinant chloroperoxidase (rCPO) was studied and compared with that of native CPO. The specific chlorination activity (47 units/nmol) of rCPO and its pH optimum (pH 2.75) were very similar to those of native CPO. rCPO catalyzes the oxidation of various substrates in comparable yields and selectivities to native CPO. Indole was oxidized to 2-oxindole with 99% selectivity and thioanisole to the corresponding R-sulfoxide (enantiomeric excess >98%). Incorporation of 18O from labeled H2 18O2 into the oxidized products was 100% in both cases.

[1]  A. Conesa,et al.  Studies on the Production of Fungal Peroxidases inAspergillus niger , 2000, Applied and Environmental Microbiology.

[2]  X Wang,et al.  Replacement of the proximal heme thiolate ligand in chloroperoxidase with a histidine residue. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[3]  M. Lehmann,et al.  Biophysical Characterization of Fungal Phytases (myo-Inositol Hexakisphosphate Phosphohydrolases): Molecular Size, Glycosylation Pattern, and Engineering of Proteolytic Resistance , 1999, Applied and Environmental Microbiology.

[4]  L. Hager,et al.  Highly Enantioselective Propargylic Hydroxylations Catalyzed by Chloroperoxidase , 1999 .

[5]  S. Ohk,et al.  Purification and properties of bacteriolytic enzymes from Bacillus licheniformis YS-1005 against Streptococcus mutans. , 1999, Bioscience, biotechnology, and biochemistry.

[6]  A. Basavapathruni,et al.  Chiral synthons via chloroperoxidase catalysis , 1998 .

[7]  A. Smith,et al.  Substrate binding and catalysis in heme peroxidases. , 1998, Current opinion in chemical biology.

[8]  R. Sheldon,et al.  Selective oxidations catalyzed by peroxidases , 1997 .

[9]  K. Ishimori,et al.  Hydrogen bond network in the distal site of peroxidases: spectroscopic properties of Asn70 --> Asp horseradish peroxidase mutant. , 1997, Biochemistry.

[10]  R. Sheldon,et al.  Chloroperoxidase catalyzed oxidations in t-butyl alcohol/water mixtures , 1997 .

[11]  Qin Zong Expression of Recombinant Chloroperoxidase , 1997 .

[12]  S. Colonna,et al.  A new enzymatic enantioselective synthesis of dialkyl sulfoxides catalysed by monooxygenases , 1997 .

[13]  K. Ishimori,et al.  Effects of Concerted Hydrogen Bonding of Distal Histidine on Active Site Structures of Horseradish Peroxidase ; Resonance Raman Studies with Asn-70 Mutants , 1996 .

[14]  H. Shoun,et al.  N-terminal processing and amino acid sequence of two isoforms of nitric oxide reductase cytochrome P450nor from Fusarium oxysporum. , 1996, Journal of biochemistry.

[15]  K. Ishimori,et al.  Catalytic roles of the distal site asparagine-histidine couple in peroxidases. , 1996, Biochemistry.

[16]  K. Ishimori,et al.  The distal glutamic acid as an acid-base catalyst in the distal site of horseradish peroxidase. , 1996, Biochemical and biophysical research communications.

[17]  P. Ortiz de Montellano,et al.  Rescue of His-42 → Ala Horseradish Peroxidase by a Phe-41 → His Mutation , 1996, The Journal of Biological Chemistry.

[18]  R. Sheldon,et al.  Synthesis of substituted oxindoles by chloroperoxidase catalyzed oxidation of indoles , 1996 .

[19]  R. Libby,et al.  Quantitating Direct Chlorine Transfer from Enzyme to Substrate in Chloroperoxidase-catalyzed Reactions* , 1996, The Journal of Biological Chemistry.

[20]  P. D. de Montellano,et al.  Rescue of the Catalytic Activity of an H42A Mutant of Horseradish Peroxidase by Exogenous Imidazoles* , 1996, The Journal of Biological Chemistry.

[21]  F. Lakner,et al.  Chloroperoxidase as Enantioselective Epoxidation Catalyst: An Efficient Synthesis of (R)-(-)-Mevalonolactone. , 1996, The Journal of organic chemistry.

[22]  Andrew T. Smith,et al.  Recombinant horseradish peroxidase isoenzyme C: the effect of distal haem cavity mutations (His42→Leu and Arg38→Leu) on compound I formation and substrate binding , 1996, JBIC Journal of Biological Inorganic Chemistry.

[23]  M. Tien,et al.  Efficient expression of a Phanerochaete chrysosporium manganese peroxidase gene in Aspergillus oryzae , 1996, Applied and environmental microbiology.

[24]  Andrew T. Smith,et al.  Role of Arginine 38 in Horseradish Peroxidase , 1996, The Journal of Biological Chemistry.

[25]  T. C. Nesbitt,et al.  Twenty-five coregulated transcripts define a sterigmatocystin gene cluster in Aspergillus nidulans. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[26]  D. Dodds,et al.  Chloroperoxidase-catalyzed asymmetric oxidations: substrate specificity and mechanistic study. , 1995 .

[27]  P. Osmulski,et al.  High-pressure-assisted reconstitution of recombinant chloroperoxidase. , 1995, Biochemistry.

[28]  T. Poulos,et al.  The crystal structure of chloroperoxidase: a heme peroxidase--cytochrome P450 functional hybrid. , 1995, Structure.

[29]  N. Kunishima,et al.  Cloning, sequencing, and heterologous expression of a gene coding for Arthromyces ramosus peroxidase. , 1995, Bioscience, biotechnology, and biochemistry.

[30]  S. Ozaki,et al.  Molecular Engineering of Horseradish Peroxidase: Thioether Sulfoxidation and Styrene Epoxidation by Phe-41 Leucine and Threonine Mutants , 1995 .

[31]  P. Ortiz de Montellano,et al.  Chloroperoxidase-catalyzed benzylic hydroxylation. , 1995, Archives of biochemistry and biophysics.

[32]  A. Dexter,et al.  Highly Enantioselective Epoxidation of 1,1-Disubstituted Alkenes Catalyzed by Chloroperoxidase , 1995 .

[33]  K. van Pée,et al.  Purification and properties of a non-haem chloroperoxidase from Serratia marcescens. , 1995, FEMS microbiology letters.

[34]  K. Hodgson,et al.  X-ray Absorption Near Edge Studies of Cytochrome P-450-CAM, Chloroperoxidase, and Myoglobin , 1995, The Journal of Biological Chemistry.

[35]  Shiro Kobayashi,et al.  Enzymatic polymerization and oligomerization , 1995 .

[36]  A. Smith,et al.  pH-dependent properties of a mutant horseradish peroxidase isoenzyme C in which Arg38 has been replaced with lysine. , 1994, European journal of biochemistry.

[37]  A. Smith,et al.  Characterization of recombinant horseradish peroxidase C and three site-directed mutants, F41V, F41W, and R38K, by resonance Raman spectroscopy. , 1994, Biochemistry.

[38]  M. Gold,et al.  Characterization of the mnp2 gene encoding manganese peroxidase isozyme 2 from the basidiomycete Phanerochaete chrysosporium. , 1994, Gene.

[39]  S. Ozaki,et al.  Molecular Engineering of Horseradish Peroxidase. Highly Enantioselective Sulfoxidation of Aryl Alkyl Sulfides by the Phe-41 .fwdarw. Leu Mutant , 1994 .

[40]  K. van Pée,et al.  Chloroperoxidase from Streptomyces lividans: isolation and characterization of the enzyme and the corresponding gene , 1994, Journal of bacteriology.

[41]  R. Sheldon,et al.  A SIMPLE PURIFICATION METHOD FOR CHLOROPEROXIDASE AND ITS USE IN ORGANIC MEDIA , 1994 .

[42]  E. Allain,et al.  Highly Enantioselective Epoxidation of Disubstituted Alkenes with Hydrogen Peroxide Catalyzed by Chloroperoxidase. , 1993 .

[43]  R. Wever,et al.  The chloroperoxidase from the fungus Curvularia inaequalis; a novel vanadium enzyme. , 1993, Biochimica et biophysica acta.

[44]  S. Colonna,et al.  Chloroperoxidase and hydrogen peroxide: An efficient system for enzymatic enantioselective sulfoxidations. , 1992 .

[45]  P. Punt,et al.  Transformation of filamentous fungi based on hygromycin B and phleomycin resistance markers. , 1992, Methods in enzymology.

[46]  J. Dordick Enzymatic and chemoenzymatic approaches to polymer synthesis. , 1992, Trends in biotechnology.

[47]  Markku Saloheimo,et al.  Heterologous Production of a Ligninolytic Enzyme: Expression of the Phlebia Radiata Laccase Gene in Trichoderma Reesei , 1991, Bio/Technology.

[48]  M. Brewster,et al.  Peroxidase-catalyzed S-oxygenation: mechanism of oxygen transfer for lactoperoxidase. , 1991, Biochemistry.

[49]  C. A. Reddy,et al.  Cloning and characterization of a lignin peroxidase gene from the white-rot fungus Trametes versicolor. , 1991, Biochemical and biophysical research communications.

[50]  A. Rasooly,et al.  Cloning of several lignin peroxidase (LIP)-encoding genes: sequence analysis of the LIP6 gene from the white-rot basidiomycete, Phanerochaete chrysosporium. , 1991, Gene.

[51]  P. Pouwels,et al.  Intracellular and extracellular production of proteins in Aspergillus under the control of expression signals of the highly expressed Aspergillus nidulans gpdA gene. , 1991, Journal of biotechnology.

[52]  M. Gold,et al.  Characterization of a gene encoding a manganese peroxidase from Phanerochaete chrysosporium. , 1990, Gene.

[53]  J. Drabowicz,et al.  A convenient procedure for the oxidation of sterically hindered sulfides to sulfoxides , 1990 .

[54]  S. Blanke,et al.  Identification of the fifth axial heme ligand of chloroperoxidase. , 1988, The Journal of biological chemistry.

[55]  J. Dawson,et al.  Cytochrome P-450 and chloroperoxidase: thiolate-ligated heme enzymes. Spectroscopic determination of their active-site structures and mechanistic implications of thiolate ligation , 1987 .

[56]  M. Nakano,et al.  On the mechanism of the peroxidase-catalyzed oxygen-transfer reaction. , 1987, Biochemistry.

[57]  L. Hager,et al.  Post-translational modifications of chloroperoxidase from Caldariomyces fumago. , 1987, Archives of biochemistry and biophysics.

[58]  J. Kelly,et al.  Transformation of Aspergillus niger by the amdS gene of Aspergillus nidulans. , 1985, The EMBO journal.

[59]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[60]  M. D. Corbett,et al.  Peroxide oxidation of indole to oxindole by chloroperoxidase catalysis. , 1979, The Biochemical journal.

[61]  J. Peisach,et al.  A model compound study of the CO-adduct of cytochrome P-450. , 1974, The Journal of biological chemistry.

[62]  D. Morris,et al.  Chloroperoxidase. I. Isolation and properties of the crystalline glycoprotein. , 1966, The Journal of biological chemistry.

[63]  G. Ellman,et al.  Tissue sulfhydryl groups. , 1959, Archives of biochemistry and biophysics.