van Pee KH

References (27)

Title : Enzymatic halogenation catalyzed via a catalytic triad and by oxidoreductases - van Pee_2000_Biol.Chem_381_1
Author(s) : van Pee KH , Keller S , Wage T , Wynands I , Schnerr H , Zehner S
Ref : Biol Chem , 381 :1 , 2000
Abstract : During the search for haloperoxidases in bacteria we detected a type of enzymes that catalyzed the peroxide-dependent halogenation of organic substrates. However, in contrast to already known haloperoxidases, these enzymes do not contain a prosthetic group or metal ions nor any other cofactor. Biochemical and molecular genetic studies revealed that they contain a catalytic triad consisting of a serine, a histidine, and an aspartate. The reaction they catalyze is actually the perhydrolysis of an acetic acid serine ester leading to the formation of peracetic acid. As a strong oxidizing agent the enzymatically formed peracetic acid can oxidize halide ions, resulting in the formation of hypohalous acid which then acts as the actual halogenating agent. Since hypohalous acid is also formed by the heme- and vanadium-containing haloperoxidases, enzymatic halogenation catalyzed by haloperoxidases and perhydrolases in general lacks substrate specificity and regioselectivity. However, detailed studies on the biosynthesis of several halometabolites led to the detection of a novel type of halogenases. These enzymes consist of a two-component system and require NADH and FAD for activity. Whereas the gene for one of the components is part of the biosynthetic cluster of the halometabolite, the second component is an enzyme which is also present in bacteria from which no halometabolites have ever been isolated, like Escherichia coli. In contrast to haloperoxidases and perhydrolases the newly detected NADH/FAD-dependent halogenases are substrate-specific and regioselective and might provide ideal tools for specific halogenation reactions.
ESTHER : van Pee_2000_Biol.Chem_381_1
PubMedSearch : van Pee_2000_Biol.Chem_381_1
PubMedID: 10722044

Title : Structural investigation of the cofactor-free chloroperoxidases - Hofmann_1998_J.Mol.Biol_279_889
Author(s) : Hofmann B , Tolzer S , Pelletier I , Altenbuchner J , van Pee KH , Hecht HJ
Ref : Journal of Molecular Biology , 279 :889 , 1998
Abstract : The structures of cofactor-free haloperoxidases from Streptomyces aureofaciens, Streptomyces lividans, and Pseudomonas fluorescens have been determined at resolutions between 1.9 A and 1.5 A. The structures of two enzymes complexed with benzoate or propionate identify the binding site for the organic acids which are required for the haloperoxidase activity. Based on these complexes and on the structure of an inactive variant, a reaction mechanism is proposed for the halogenation reaction with peroxoacid and hypohalous acid as reaction intermediates. Comparison of the structures suggests that a specific halide binding site is absent in the enzymes but that hydrophobic organic compounds may fit into the active site pocket for halogenation at preferential sites.
ESTHER : Hofmann_1998_J.Mol.Biol_279_889
PubMedSearch : Hofmann_1998_J.Mol.Biol_279_889
PubMedID: 9642069
Gene_locus related to this paper: psefl-cpoF , strau-brpa2 , strau-cpoT , strli-cpoL

Title : Regulation of activity of chloroperoxidase from Serratia marcescens - Burd_1998_Biochemistry_63_1299
Author(s) : Burd VN , Vasilyeva OV , Voskoboev AI , van Pee KH
Ref : Biochemistry , 63 :1299 , 1998
Abstract : The influence of various factors on the activity of chloroperoxidase from Serratia marcescens was investigated. The enzyme is active only in acetate-containing buffers within the pH range 4.2-5.8. F-, Cu2+, [Fe(CN)6]4+, and [Fe(CN)6]3+ inhibit the enzyme. The chloroperoxidase is thermostable and resistant to the effect of lower alcohols.
ESTHER : Burd_1998_Biochemistry_63_1299
PubMedSearch : Burd_1998_Biochemistry_63_1299
PubMedID: 9864470

Title : Thiocarbamate herbicide-inducible nonheme haloperoxidase of Rhodococcus erythropolis NI86\/21 - De Schrijver_1997_Appl.Environ.Microbiol_63_1911
Author(s) : De Schrijver A , Nagy I , Schoofs G , Proost P , Vanderleyden J , van Pee KH , De Mot R
Ref : Applied Environmental Microbiology , 63 :1911 , 1997
Abstract : During biodegradation of thiocarbamate herbicides by Rhodococcus erythropolis NI86/21, a protein with an M(r) of 30,000 is induced (I. Nagy, G. Schoofs, F. Compernolle, P. Proost, J. Vanderleyden, and R.De Mot, J. Bacteriol. 177:676-687, 1995). Based on N-terminal sequence data for the protein purified by two-dimensional electrophoresis, the corresponding structural gene, thcF, was cloned and sequenced. The deduced protein sequence of ThcF is homologous to those of nonheme haloperoxidases. A particularly high level of sequence identity (72.6%) was observed for the chloroperoxidase from Pseudomonas pyrrocinia. A polyclonal antibody against the latter enzyme cross-reacted with ThcF either produced by the original Rhodococcus cells or overexpressed heterologously in Escherichia coli. In both thiocarbamate-grown Rhodococcus cells and E. coli cells expressing thcF, the haloperoxidase activity of ThcF was demonstrated. The thiocarbamate-inducible R. erythropolis ThcF protein represents the first (nonheme) haloperoxidase to be identified in a nocardioform actinomycete.
ESTHER : De Schrijver_1997_Appl.Environ.Microbiol_63_1911
PubMedSearch : De Schrijver_1997_Appl.Environ.Microbiol_63_1911
PubMedID: 9143122
Gene_locus related to this paper: rhoer-thcf

Title : The non-haem chloroperoxidase from Pseudomonas fluorescens and its relationship to pyrrolnitrin biosynthesis - Kirner_1996_Microbiology_142_2129
Author(s) : Kirner S , Krauss S , Sury G , Lam ST , Ligon JM , van Pee KH
Ref : Microbiology , 142 :2129 , 1996
Abstract : The non-haem chloroperoxidase gene (cpoF) from the pyrrolnitrin producer Pseudomonas fluorescens BL914 was cloned using an oligonucleotide derived from part of the N-terminal amino acid sequence of chloroperoxidase (CPO-P) from Pseudomonas pyrrocina as a probe. Based on the overexpression of cpoF in Escherichia coli and the stability of CPO-F against higher temperatures and proteases, the enzyme was purified to homogeneity. Partial characterization of the enzyme showed that it belongs to the class of bacterial non-haem CPOs. To investigate the role of CPO-F in pyrrolnitrin biosynthesis, the cpoF gene was inactivated by insertion of a kanamycin cassette. Exchange of the chromosomal cpoF gene against the disrupted copy had no influence on pyrrolnitrin production demonstrating that CPO-F was not involved in pyrrolnitrin biosynthesis.
ESTHER : Kirner_1996_Microbiology_142_2129
PubMedSearch : Kirner_1996_Microbiology_142_2129
PubMedID: 8760926
Gene_locus related to this paper: psefl-cpoF

Title : Cloning, sequencing and disruption of a bromoperoxidase-catalase gene in Streptomyces venezuelae: evidence that it is not required for chlorination in chloramphenicol biosynthesis - Facey_1996_Microbiology_142_657
Author(s) : Facey SJ , Gross F , Vining LC , Yang K , van Pee KH
Ref : Microbiology , 142 :657 , 1996
Abstract : Genomic DNA libraries of Streptomyces venezuelae ISP5230 and of a mutant blocked at the chlorination step of chloramphenicol biosynthesis were probed by hybridization with a synthetic oligonucleotide corresponding to the N-terminal amino acid sequence of a bromoperoxidase-catalase purified from the wild-type strain. Hybridizing fragments obtained from the two strains were cloned and sequenced. Analysis of the nucleotide sequences demonstrated that the fragments contained the same 1449 bp open reading frame with no differences in nucleotide sequence. The deduced polypeptide encoded 483 amino acids with a calculated M(r) of 54,200; the N-terminal sequence was identical to that of the bromoperoxidase-catalase purified from wild-type S. venezuelae. Comparison of the amino acid sequence predicted for the cloned bromoperoxidase-catalase gene (bca) with database protein sequences showed a significant similarity to a group of prokaryotic and eukaryotic catalases, but none to other peroxidases or haloperoxidases. Replacement of the bca gene in the wild-type strain of S. venezuelae with a copy disrupted by insertion of a DNA fragment encoding apramycin resistance did not prevent chloramphenicol production. The results suggest that the role of the enzyme in S. venezuelae is related to its activity as a catalase rather than as a halogenating agent.
ESTHER : Facey_1996_Microbiology_142_657
PubMedSearch : Facey_1996_Microbiology_142_657
PubMedID: 8868441

Title : Biosynthesis of halogenated metabolites by bacteria - van Pee_1996_Annu.Rev.Microbiol_50_375
Author(s) : van Pee KH
Ref : Annu Rev Microbiol , 50 :375 , 1996
Abstract : Halogenated metabolites, originally thought to be infrequent in nature, are actually nothing unusual at all, and are produced by many different organisms, including bacteria. Whereas marine bacteria usually produce brominated compounds, terrestrial bacteria preferentially synthesize chlorometabolites, but fluoro- and iodometabolites can also be found. Haloperoxidases, enzymes capable of catalyzing the formation of carbon halogen bonds in the presence of hydrogen peroxide and halide ions (Cl-, Br- and I-) have been isolated and characterized from different bacteria. These enzymes turned out to be very unspecific and are obviously not the type of halogenating enzymes responsible for the formation of halometabolites in bacteria. A yet-unknown type of halogenating enzyme having both substrate and regio-specificity must be involved in the biosynthesis of halogenated compounds.
ESTHER : van Pee_1996_Annu.Rev.Microbiol_50_375
PubMedSearch : van Pee_1996_Annu.Rev.Microbiol_50_375
PubMedID: 8905085

Title : Purification and properties of a non-haem chloroperoxidase from Serratia marcescens - Burd_1995_FEMS.Microbiol.Lett_129_255
Author(s) : Burd W , Yourkevich O , Voskoboev AJ , van Pee KH
Ref : FEMS Microbiology Letters , 129 :255 , 1995
Abstract : A non-haem chloroperoxidase was isolated from the enteric bacterium Serratia marcescens. The enzyme was purified to homogeneity by heat treatment, ammonium sulfate precipitation, ion exchange chromatography, gel filtration and dye-ligand affinity chromatography. Native chloroperoxidase has a molecular mass of 58 kDa and consists of two identical subunits of 29 kDa. Whereas chloroperoxidase catalyses only the bromination of monochlorodimedone, indole is chlorinated by this enzyme. Chloroperoxidase also catalyses the oxidation of amino to nitro groups. The enzyme is thermostable and does not lose any activity when incubated at 65 degrees C for 2 h. Comparison of the first 15 amino-terminal amino acids showed a sequence identity of 80% to the chloroperoxidases from Streptomyces lividans and Pseudomonas pyrrocinia. However, no precipitation band was obtained in the Ouchterlony agar diffusion assay with antibodies raised against the chloroperoxidases from Pseudomonas pyrrocinia and Streptomyces aureofaciens Tu24.
ESTHER : Burd_1995_FEMS.Microbiol.Lett_129_255
PubMedSearch : Burd_1995_FEMS.Microbiol.Lett_129_255
PubMedID: 7607409

Title : Chloroperoxidase from Streptomyces lividans: isolation and characterization of the enzyme and the corresponding gene - Bantleon_1994_J.Bacteriol_176_2339
Author(s) : Bantleon R , Altenbuchner J , van Pee KH
Ref : Journal of Bacteriology , 176 :2339 , 1994
Abstract : For the first time, a halogenating enzyme which is not known to produce halogenated metabolites has been isolated from a bacterial strain. The gene encoding the nonheme chloroperoxidase (CPO-L) from Streptomyces lividans TK64 was cloned, and its gene product was characterized. S. lividans TK64 produced only very small amounts of the enzyme. After cloning of the gene into Streptomyces aureofaciens T24-88, the enzyme was overexpressed up to 3,000-fold. Based on the overexpression, a simple purification procedure using acid precipitation and hydrophobic interaction chromatography was developed. Thus, 54 mg of homogeneous CPO-L could be obtained from 27 g (wet weight) of mycelium. The native enzyme has a molecular weight of 64,000 and consists of two identical subunits. The enzyme does not exhibit an absorption peak in the Soret region of the optical spectrum. X-ray fluorescence spectroscopy revealed that the enzyme does not contain any metal ions in equimolar amounts. CPO-L showed cross-reaction with antibodies raised against the nonheme chloroperoxidase from Pseudomonas pyrrocinia but not with antibodies raised against CPO-T from S. aureofaciens T24. CPO-L exhibits substrate specificity only for chlorination, not for bromination. Therefore, monochlorodimedone is only brominated by CPO-L, whereas indole is brominated and chlorinated. The functional chloroperoxidase gene was located on a 1.9-kb SalI DNA fragment. DNA sequence analysis revealed an open reading frame encoding a predicted polypeptide of 276 amino acids. The overall identity of the amino acid sequence to that of chloroperoxidase from P. pyrrocinia was 71%, whereas that to bromoperoxidase BPO-A2 from S. aureofaciens ATCC 10762 was only 42%
ESTHER : Bantleon_1994_J.Bacteriol_176_2339
PubMedSearch : Bantleon_1994_J.Bacteriol_176_2339
PubMedID: 8157602
Gene_locus related to this paper: strli-cpoL

Title : Cloning of a second non-haem bromoperoxidase gene from Streptomyces aureofaciens ATCC 10762: sequence analysis, expression in Streptomyces lividans and enzyme purification - Pelletier_1994_Microbiology_140_509
Author(s) : Pelletier I , Pfeifer O , Altenbuchner J , van Pee KH
Ref : Microbiology , 140 :509 , 1994
Abstract : The gene for BPO-A1, one of two non-haem bromoperoxidases in the tetracycline and 7-chlorotetracycline producer Streptomyces aureofaciens ATCC 10762, was cloned in the positive selection vector pIJ699 and expressed in Streptomyces lividans TK64. The cloned bromoperoxidase was over-produced up to 2800-fold by the S. lividans TK64 transformant. By taking advantage of the over-production of BPO-A1 and the heat stability of the enzyme, a new and simple purification procedure was developed. Subcloning into the vector pIJ487 and screening of recombinants by a newly developed histochemical assay located the bpoA1 gene on a 2.1 kb BamHI-HindIII fragment. The nucleotide sequence of the 2.1 kb fragment was determined; the bpoA1 gene was identified within the sequence on the basis of the biased codon usage of Streptomyces genes and the presence of a nucleotide sequence encoding the N-terminal amino acid sequence obtained from the purified BPO-A1. Comparison of the deduced primary structure of BPO-A1 with those deduced for the non-haem chloroperoxidase CPO-P from Pseudomonas pyrrocinia and the bromoperoxidase BPO-A2 from S. aureofaciens ATCC 10762 gave amino acid sequence identities of 49% and 40%, respectively.
ESTHER : Pelletier_1994_Microbiology_140_509
PubMedSearch : Pelletier_1994_Microbiology_140_509
PubMedID: 8012573
Gene_locus related to this paper: strau-brpa1

Title : The metal-ion-free oxidoreductase from Streptomyces aureofaciens has an alpha\/beta hydrolase fold - Hecht_1994_Nat.Struct.Biol_1_532
Author(s) : Hecht HJ , Sobek H , Haag T , Pfeifer O , van Pee KH
Ref : Nat Struct Biol , 1 :532 , 1994
Abstract : The crystal structure of the bromoperoxidase A2 from Streptomyces aureofaciens (ATCC 10762) has been determined by isomorphous replacement and refined to 2.05 A resolution with an R-value of 18.4%. The enzyme catalyzes the bromination of organic compounds in the presence of bromide and peroxide. The structure confirms the absence of cofactors such as metal ions or haem groups and shows the general topology of the alpha/beta hydrolase fold. The active centre is at the end of a deep pocket and includes a catalytic triad of Ser 98, Asp 228 and His 257. The active centre is connected by a narrow tunnel to a second pocket on the enzyme surface.
ESTHER : Hecht_1994_Nat.Struct.Biol_1_532
PubMedSearch : Hecht_1994_Nat.Struct.Biol_1_532
PubMedID: 7664081
Gene_locus related to this paper: strau-brpa2

Title : The catechol 2,3-dioxygenase gene of Rhodococcus rhodochrous CTM: nucleotide sequence, comparison with isofunctional dioxygenases and evidence for an active-site histidine - Candidus_1994_Microbiology_140_321
Author(s) : Candidus S , van Pee KH , Lingens F
Ref : Microbiology , 140 :321 , 1994
Abstract : In cell-free extracts of Escherichia coli clones harbouring the 3.5 kb Bg/II fragment of plasmid pTC1 from Rhodococcus rhodochrous CTM a catechol 2,3-dioxygenase (C23O) accepting both 3-methylcatechol and 2,3-dihydroxybiphenyl as substrates could be detected. The plasmid-encoded gene for C23O of R. rhodochrous CTM and its flanking regions were sequenced. In front of the gene a sequence resembling an E. coli promoter was identified, which led to constitutive expression of the cloned gene in E. coli TG1. The derived amino acid sequence of the C23O was compared to that of nine other enzymes, which all catalyse the extradiol cleavage of an aromatic ring. These nine sequences were from different Pseudomonas strains, in contrast to the sequence described here, from a Gram-positive bacterium. The role of four strongly conserved histidines was examined by chemical modification of the histidyl residues of the native enzyme by diethylpyrocarbonate. For that purpose the C23O was purified to homogeneity from E. coli harbouring pSC1701. However, the enzyme lost its activity during the purification. Activity could partially be restored by treatment with Fe2+ and reducing agents.
ESTHER : Candidus_1994_Microbiology_140_321
PubMedSearch : Candidus_1994_Microbiology_140_321
PubMedID: 8180697

Title : Chloroperoxidase-encoding gene from Pseudomonas pyrrocinia: sequence, expression in heterologous hosts, and purification of the enzyme - Wolfframm_1993_Gene_130_131
Author(s) : Wolfframm C , Lingens F , Mutzel R , van Pee KH
Ref : Gene , 130 :131 , 1993
Abstract : The nucleotide sequence of a 1.5-kb fragment of Pseudomonas pyrrocinia DNA containing the chloroperoxidase(CPO)-encoding gene (cpo) and its flanking regions was determined. The cpo codes for a protein of 278 amino acids (aa). The mature enzyme contains no N-terminal methionine, so that the CPO monomer consists of 277 aa with a calculated M(r) of 30,304. Expression studies showed that the cpo from P. pyrrocinia is functionally expressed in Escherichia coli and Streptomyces lividans. Based on the overproduction of the CPO in E. coli, a novel and simple purification procedure was developed allowing the isolation of about 800-fold more CPO per gram of cells than was originally isolated from P. pyrrocinia. Comparison with the aa sequence of the bromoperoxidase BPO-A2 from S. aureofaciens ATCC10762 revealed an identity of 38%.
ESTHER : Wolfframm_1993_Gene_130_131
PubMedSearch : Wolfframm_1993_Gene_130_131
PubMedID: 8344520
Gene_locus related to this paper: psepy-prxc

Title : Molecular cloning and sequencing of a non-haem bromoperoxidase gene from Streptomyces aureofaciens ATCC 10762 - Pfeifer_1992_J.Gen.Microbiol_138_1123
Author(s) : Pfeifer O , Pelletier I , Altenbuchner J , van Pee KH
Ref : J Gen Microbiol , 138 :1123 , 1992
Abstract : A bromoperoxidase gene (bpoT), recently cloned from Streptomyces aureofaciens Tu24, was used as a probe in Southern blot hybridization of total DNA from S. aureofaciens ATCC 10762. A single SstI fragment of 5.4 kb was detected, which was cloned via an enriched gene library into Escherichia coli. The functional bromoperoxidase gene was located on a 2.1 kb BamHI-HindIII fragment by subcloning into S. lividans TK64, using the multicopy plasmid pIJ486. The enzyme was overproduced in S. lividans TK64 (up to 30,000 times compared to S. aureofaciens ATCC 10762) and showed the same electrophoretic and immunological properties as the bromoperoxidase BPO-A2 purified from S. aureofaciens ATCC 10762. DNA sequence analysis revealed an open reading frame encoding a predicted polypeptide with the same M(r) and N-terminal amino acid sequence as the purified subunit of BPO-A2.
ESTHER : Pfeifer_1992_J.Gen.Microbiol_138_1123
PubMedSearch : Pfeifer_1992_J.Gen.Microbiol_138_1123
PubMedID: 1527491
Gene_locus related to this paper: strau-brpa2

Title : Purification, characterization and comparison of two non-haem bromoperoxidases from Streptomyces aureofaciens ATCC 10762 - Weng_1991_J.Gen.Microbiol_137_2539
Author(s) : Weng M , Pfeifer O , Krauss S , Lingens F , van Pee KH
Ref : J Gen Microbiol , 137 :2539 , 1991
Abstract : Two non-haem bromoperoxidases (BPO 1 and BPO 2) were purified from the 7-chlorotetracycline-producing strain Streptomyces aureofaciens ATCC 10762. Both enzymes showed azide-insensitive brominating activity, and bromide-dependent peroxidase activity. BPO 1 was a dimer (Mr 65,000) with subunits of identical size (Mr 31,000). The pI was estimated to be 4.5. The enzyme did not cross-react with antibodies raised against the non-haem bromoperoxidase (Mr 90,000) from S. aureofaciens Tu24, a strain that also produces 7-chlorotetracycline. The Mr of BPO 2 was estimated to be 90,000. The enzyme had three identical subunits (Mr 31,000), and its isoelectric point was 3.5, identical with that of the bromoperoxidase from S. aureofaciens Tu24. Moreover, BPO 2 was immunologically identical with the bromoperoxidase from S. aureofaciens Tu24, although both it and BPO 1 could be distinguished electrophoretically from the latter bromoperoxidase.
ESTHER : Weng_1991_J.Gen.Microbiol_137_2539
PubMedSearch : Weng_1991_J.Gen.Microbiol_137_2539
PubMedID: 1783900
Gene_locus related to this paper: strau-brpa1 , strau-brpa2

Title : Crystallization and preliminary X-ray data of bromoperoxidase from Streptomyces aureofaciens ATCC 10762 - Sobek_1991_J.Mol.Biol_221_35
Author(s) : Sobek H , Haag T , Pfeifer O , Schomburg D , Lingens F , van Pee KH
Ref : Journal of Molecular Biology , 221 :35 , 1991
Abstract : Bromoperoxidase from Streptomyces aureofaciens ATCC 10762, a non-haem haloperoxidase, has been crystallized using the hanging drop method. Preliminary X-ray diffraction studies show that the crystals belong to the cubic space group P2(1)3 with a = 123.4 A. The asymmetric unit contains a dimer of Mr = 60,200. The crystals diffract to at least 2.3 A resolution and are suitable for crystallographic structure analysis.
ESTHER : Sobek_1991_J.Mol.Biol_221_35
PubMedSearch : Sobek_1991_J.Mol.Biol_221_35
PubMedID: 1920414

Title : Purification, properties and immunological detection of a bromoperoxidase-catalase from Streptomyces venezuelae and from a chloramphenicol-nonproducing mutant - Knoch_1989_J.Gen.Microbiol_135_2493
Author(s) : Knoch M , van Pee KH , Vining LC , Lingens F
Ref : J Gen Microbiol , 135 :2493 , 1989
Abstract : A new bromoperoxidase-catalase was purified from the chloramphenicol-producing actinomycete Streptomyces venezuelae ISP 5230. The homogeneous enzyme showed brominating activity, catalase activity and a very low peroxidase activity. The spectral properties and pH dependence of the catalase activity showed similarities to conventional catalases. In contrast to other haem-bromoperoxidases, the bromoperoxidase-catalase was stable when treated with an ethanol/chloroform mixture. Gel filtration gave an estimated Mr of 127,000-136,000. SDS-PAGE showed a single band corresponding in mobility to a species with an Mr of 61,000. The pI was estimated to be 4.5. The bromoperoxidase-catalase was not present in active form in a mutant of S. venezuelae ISP 5230, blocked in the chlorination step of chloramphenicol biosynthesis. However, an inactive species of the enzyme was detected in crude extracts of the mutant by using antibodies. From these results it is concluded that this bromoperoxidase participates in the chlorination step during chloramphenicol biosynthesis.
ESTHER : Knoch_1989_J.Gen.Microbiol_135_2493
PubMedSearch : Knoch_1989_J.Gen.Microbiol_135_2493
PubMedID: 2628543

Title : Cloning and high-level expression of a chloroperoxidase gene from Pseudomonas pyrrocinia in Escherichia coli - Wolfframm_1988_FEBS.Lett_238_325
Author(s) : Wolfframm C , van Pee KH , Lingens F
Ref : FEBS Letters , 238 :325 , 1988
Abstract : A chloroperoxidase gene from Pseudomonas pyrrocinia was cloned into Escherichia coli using the cosmid vector pJB8. The gene coding for the chloroperoxidase could be localized to a 1.5 kb fragment of DNA which was subcloned into the high-copy-number plasmid pUC18. In one subclone increased halogenating activity could be found which was 570-fold greater than in P. pyrrocinia. The halogenating enzyme was identified as the chloroperoxidase by SDS-polyacrylamide gel electrophoresis.
ESTHER : Wolfframm_1988_FEBS.Lett_238_325
PubMedSearch : Wolfframm_1988_FEBS.Lett_238_325
PubMedID: 3049160

Title : Molecular cloning and high-level expression of a bromoperoxidase gene from Streptomyces aureofaciens Tu24 - van Pee_1988_J.Bacteriol_170_5890
Author(s) : van Pee KH
Ref : Journal of Bacteriology , 170 :5890 , 1988
Abstract : A bromoperoxidase gene was cloned from Streptomyces aureofaciens T24 into Streptomyces lividans TK64 by using the promoter-probe vector pIJ486. Subcloning of DNA from the original, unstable clone allowed the gene to be localized to a 1.7-kilobase (kb) fragment of DNA. Southern blotting showed that the cloned 1.7-kb insert hybridized to a 4.3-kb fragment in an SstI digest of S. aureofaciens T24 total DNA. The 1.7-kb insert was shown to code for a protein with the electrophoretic properties of the subunits of the nonheme bromoperoxidase isolated from S. aureofaciens T24. The protein produced by S. lividans TK64 transformed with pHM621, which contained an 8.0-kb insert, was shown to be identical to the S. aureofaciens T24 bromoperoxidase in terms of its electrophoretic mobility on denaturing and nondenaturing polyacrylamide gels and its NH2-terminal amino acid sequence. The bromoperoxidase was overproduced (up to 180 times) by S. lividans TK64 containing pHM621. Based on the heat stability of the S. aureofaciens T24 bromoperoxidase, a new and simple purification procedure with very high yields was developed.
ESTHER : van Pee_1988_J.Bacteriol_170_5890
PubMedSearch : van Pee_1988_J.Bacteriol_170_5890
PubMedID: 3142859

Title : Purification and partial characterization of multiple bromoperoxidases from Streptomyces griseus - Zeiner_1988_J.Gen.Microbiol_134_3141
Author(s) : Zeiner R , van Pee KH , Lingens F
Ref : J Gen Microbiol , 134 :3141 , 1988
Abstract : The presence of multiple bromoperoxidases in extracts of Streptomyces griseus T 6 was detected. The enzyme pattern varied with the age of the culture. A haem-type bromoperoxidase (BPO 2) was always present. Additionally three nonhaem-type bromoperoxidases (BPO 1a, 1b and 3) were detected and purified to homogeneity. The Mr of non-denatured BPO 1a was 70,000 +/- 10,000 and those of BPO 1b and 3 were 90,000 +/- 5000. BPO 1a and 1b were dimers with subunit Mr values of 34,000 and 43,000, respectively. BPO 3 was a trimer with a subunit Mr of 31,000. The enzymes differed in their isoelectric points, heat stability, and Km values. In immunodiffusion experiments BPO 1a and 3 showed partial identity with the nonhaem-type bromoperoxidase from Streptomyces aureofaciens. The nonhaem-type BPO 1a, 1b and 3 had neither peroxidase nor catalase activity.
ESTHER : Zeiner_1988_J.Gen.Microbiol_134_3141
PubMedSearch : Zeiner_1988_J.Gen.Microbiol_134_3141
PubMedID: 3151989

Title : Purification and characterization of a novel bacterial non-heme chloroperoxidase from Pseudomonas pyrrocinia - Wiesner_1988_J.Biol.Chem_263_13725
Author(s) : Wiesner W , van Pee KH , Lingens F
Ref : Journal of Biological Chemistry , 263 :13725 , 1988
Abstract : The first bacterial chloroperoxidase that is capable of catalyzing the chlorination of indole to 7-chloroindole was detected in Pseudomonas pyrrocinia ATCC 15958, a bacterium that produces the antifungal antibiotic pyrrolnitrin (Wiesner, W., van Pe, K.H., and Lingens, F. (1986) FEBS Lett. 209, 321-324). Here we describe the purification and characterization of this bacterial non-heme chloroperoxidase. The enzyme was purified by DEAE-cellulose chromatography at different pH values, molecular sieve chromatography, and Bio-Gel HTP hydroxylapatite. After the last purification step, chloroperoxidase was homogeneous by polyacrylamide gel electrophoresis and ultracentrifugation. Based on gel filtration and ultracentrifugation results, the molecular weight of the enzyme was 64,000 +/- 3,000. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed a single band with the mobility of a 32,000 molecular weight species. Therefore, in solution at neutral pH, this chloroperoxidase is a dimer. The enzyme did not exhibit any absorbance in the visible region of the spectrum. The isoelectric point was 4.1. Chloroperoxidase was specific for I-, Br-, and Cl- and was not inhibited by azide, but was inhibited by cyanide and F-. This procaryotic chloroperoxidase catalyzed the bromination of monochlorodimedone but not its chlorination and has no peroxidase or catalase activity. The pH optimum of the enzyme was between 4.0 and 4.5, and the enzyme was stable between pH 3.5 and 8.5 and showed no loss of activity when incubated at 60 degrees C for 2 h. Chloroperoxidase also chlorinated 4-(2-amino-3-chlorophenyl) pyrrole to yield aminopyrrolnitrin, the immediate precursor of pyrrolnitrin. This suggests very strongly that chloroperoxidase is involved in the biosynthesis of the antibiotic pyrrolnitrin.
ESTHER : Wiesner_1988_J.Biol.Chem_263_13725
PubMedSearch : Wiesner_1988_J.Biol.Chem_263_13725
PubMedID: 3417677

Title : Purification and properties of a nonheme bromoperoxidase from Streptomyces aureofaciens - van Pee_1987_Biol.Chem.Hoppe.Seyler_368_1225
Author(s) : van Pee KH , Sury G , Lingens F
Ref : Biol Chem Hoppe Seyler , 368 :1225 , 1987
Abstract : The first bacterial nonheme type bromoperoxidase has been purified to homogeneity from the chlorotetracycline-producing actinomycete Streptomyces aureofaciens T 24. Purification was accomplished by (NH4)2SO4 precipitation, DEAE-cellulose chromatography at different pH-values, and molecular sieve chromatography. The purified enzyme has a molecular mass of 90 to 95 kDa based on ultracentrifugation and gel filtration. The enzyme is composed of three subunits of identical molecular mass (m = 31 kDa). Bromoperoxidase catalyses the bromination of monochlorodimedone, but not its chlorination, and has no peroxidase or catalase activity. The optimum pH is 4.5. The enzyme does not exhibit an absorption peak in the Soret region of the optical spectrum. X-ray fluorescence spectroscopy revealed that the enzyme does not contain any metals in equimolar amounts. Bromoperoxidase is stable in a pH range from pH 4.0 to pH 10.0 at 4 degrees C for weeks and does not loose any activity when incubated at 80 degrees C for 2 h.
ESTHER : van Pee_1987_Biol.Chem.Hoppe.Seyler_368_1225
PubMedSearch : van Pee_1987_Biol.Chem.Hoppe.Seyler_368_1225
PubMedID: 3118902

Title : Detection of a new chloroperoxidase in Pseudomonas pyrrocinia - Wiesner_1986_FEBS.Lett_209_321
Author(s) : Wiesner W , van Pee KH , Lingens F
Ref : FEBS Letters , 209 :321 , 1986
Abstract : A new chloroperoxidase could be detected in Pseudomonas pyrrocinia ATCC 15,958, a bacterium that produces the antifungal antibiotic pyrrolnitrin. This enzyme was separated from a ferriprotoporphyrin IX containing bromoperoxidase which was also produced by this bacterium. The enzyme is capable of catalyzing the chorination of indole to 7-chloroindole. This procaryotic chloroperoxidase requires the presence of H2O2 and can also brominate monochlorodimedone, but cannot catalyze its chlorination. This enzyme is the first chloroperoxidase described from procaryotic sources.
ESTHER : Wiesner_1986_FEBS.Lett_209_321
PubMedSearch : Wiesner_1986_FEBS.Lett_209_321
PubMedID: 3792552

Title : Purification of bromoperoxidase from Pseudomonas aureofaciens - van Pee_1985_J.Bacteriol_161_1171
Author(s) : van Pee KH , Lingens F
Ref : Journal of Bacteriology , 161 :1171 , 1985
Abstract : A Bromoperoxidase has been isolated and purified from Pseudomonas aureofaciens ATCC 15926 mutant strain ACN. The purified enzyme was homogeneous as determined by polyacrylamide gel electrophoresis and ultracentrifugation. This bromoperoxidase can utilize bromide ions in the presence of hydrogen peroxide and a halogen acceptor for the catalytic formation of carbon-halogen bonds. The homogeneous enzyme also has peroxidase and catalase activity. Based on the results from gel filtration and ultracentrifugation, the molecular weight of this procaryotic bromoperoxidase is 155,000 to 158,000. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis shows a single band having the mobility of a 77,000-molecular-weight species. We thus conclude that this bromoperoxidase exists in solution as a dimeric species. The heme prosthetic group of bromoperoxidase is ferriprotoporphyrin IX. The spectral properties of the native and reduced enzyme are reported. This bromoperoxidase is the first halogenating enzyme purified from procaryotic sources.
ESTHER : van Pee_1985_J.Bacteriol_161_1171
PubMedSearch : van Pee_1985_J.Bacteriol_161_1171
PubMedID: 3972772

Title : Purification and molecular and catalytic properties of bromoperoxidase from Streptomyces phaeochromogenes - van Pee_1985_J.Gen.Microbiol_131_1911
Author(s) : van Pee KH , Lingens F
Ref : J Gen Microbiol , 131 :1911 , 1985
Abstract : A bromoperoxidase has been isolated and purified from the chloramphenicol-producing actinomycete Streptomyces phaeochromogenes. The purified enzyme was homogeneous as determined by polyacrylamide gel electrophoresis. The prosthetic group of the bromoperoxidase was ferriprotoporphyrin IX. Based on gel filtration results the molecular weight of the enzyme was 147 000 +/- 3000. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis showed a single band having the mobility of a 72 500 molecular weight species. Therefore, in solution at neutral pH, the bromoperoxidase behaved as a dimer. The isoelectric point was 4.0. The spectral properties of the native and reduced enzyme are reported. The homogeneous enzyme also had peroxidase and catalase activity.
ESTHER : van Pee_1985_J.Gen.Microbiol_131_1911
PubMedSearch : van Pee_1985_J.Gen.Microbiol_131_1911
PubMedID: 4056738

Title : Purification and properties of bromoperoxidase from Pseudomonas pyrrocinia - Wiesner_1985_Biol.Chem.Hoppe.Seyler_366_1085
Author(s) : Wiesner W , van Pee KH , Lingens F
Ref : Biol Chem Hoppe Seyler , 366 :1085 , 1985
Abstract : A bromoperoxidase was purified and partially characterized from Pseudomonas pyrrocinia ATCC 15958, a bacterium that produces the antifungal antibiotic pyrrolnitrin. The purified enzyme preparation was homogeneous as determined by polyacrylamide gel electrophoresis and ultracentrifugation. The molecular mass of the enzyme was estimated to be 154 kDa +/- 3 kDa as determined by gel filtration and ultracentrifugation. Sodium dodecyl sulfate polyacrylamide gel electrophoresis showed a single band with the mobility of a 76-kDa species. Therefore, in solution at neutral pH, bromoperoxidase exists as a dimeric species. The isoelectric point was 5.0. The prosthetic group of this procaryotic bromoperoxidase was ferriprotoporphyrin IX. The spectral properties of the native and reduced enzyme are reported. The purified enzyme showed brominating as well as peroxidase and catalase activity.
ESTHER : Wiesner_1985_Biol.Chem.Hoppe.Seyler_366_1085
PubMedSearch : Wiesner_1985_Biol.Chem.Hoppe.Seyler_366_1085
PubMedID: 4091967

Title : Detection of a bromoperoxidase in Streptomyces phaeochromogenes - van Pee_1984_FEBS.Lett_173_5
Author(s) : van Pee KH , Lingens F
Ref : FEBS Letters , 173 :5 , 1984
Abstract : A bromoperoxidase could be detected after fractionation in the chloramphenicol producing actinomycete, Streptomyces phaeochromogenes. This enzyme is capable of catalyzing the bromination of the antifungal antibiotic pyrrolnitrin [3-chloro-4-(2-nitro-3-chlorophenyl)pyrrole] in the 2-position of the pyrrole ring. The enzyme had a pH optimum of 5.0. This procaryotic bromoperoxidase requires the presence of H2O2 and can also brominate monochlorodimedone, but cannot catalyze chlorination. This enzyme is the first haloperoxidase described from procaryotic sources.
ESTHER : van Pee_1984_FEBS.Lett_173_5
PubMedSearch : van Pee_1984_FEBS.Lett_173_5
PubMedID: 6745436