Identification of Novel Non-Metal Haloperoxidases from the Marine Metagenome

Hui-Jeong  Gwon; Ide  Teruhiko; Harayama  Shigeaki  and Sang-Ho  Baik

doi:10.4014/jmb.1310.10070

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Environmental Microbiology and Engineering

Identification of Novel Non-Metal Haloperoxidases from the Marine Metagenome

Hui-Jeong Gwon ¹, Ide Teruhiko ², Harayama Shigeaki ² and Sang-Ho Baik ^{2, 3*}

¹Radiation Research Division for Industry & Environment, Korea Atomic Energy Research Institute, Jeongeup 580-185, Republic of Korea, ²Marine Biotechnology Institute, Heita, Kamaishi, Iwate 026-0001, Japan, ³Department of Food Science and Human Nutrition, and Research Institute of Makgeolli, Chonbuk National University, Jeonju 561-756, Republic of Korea

Received: October 21, 2013; Accepted: February 26, 2014

J. Microbiol. Biotechnol. 2014; 24(6): 835-842

Published June 28, 2014 https://doi.org/10.4014/jmb.1310.10070

Abstract

Haloperoxidase (HPO, E.C.1.11.1.7) is a metal-containing enzyme oxidizing halonium species, which can be used in the synthesis of halogenated organic compounds, for instance in the production of antimicrobial agents, cosmetics, etc., in the presence of halides and H2O2. To isolate and evaluate a novel non-metal HPO using a culture-independent method, a cassette PCR library was constructed from marine seawater in Japan. We first isolated a novel HPO gene from Pseudomonas putida ATCC11172 by PCR for constructing the chimeric HPO library (HPO11172). HPO11172 showed each single open-reading frame of 828 base pairs coding for 276 amino acids, respectively, and showed 87% similarity with P. putida IF-3 sequences. Approximately 600 transformants screened for chimeric genes between P. putida ATCC11173 and HPO central fragments were able to identify 113 active clones. Among them, we finally isolated 20 novel HPO genes. Sequence analyses of the obtained 20 clones showed higher homology genes with P. putida or Sinorhizobium or Streptomyces strains. Although the HPO A9 clone showed the lowest homology with HPO11172, clones in group B, including CS19, showed a relatively higher homology of 80%, with 70% identy. E. coli cells expressing these HPO chimeric genes were able to successfully bioconvert chlorodimedone with KBr or KCl as substrate.

Keywords

haloperoxidase, marine metagenome, chimeric library, cassette PCR

References

Almeida M, Filipe S, Humanes M, Maia MF, Melo R, Severino N, et al. 2001. Vanadium haloperoxidases from brown algae of the Laminariaceae family. Phytochemistry 57:633-642.
Iffland A, Gendreizig S, Tafelmeyer P, Johnsson K. 2001. Changing the substrate specificity of cytochrome c peroxidase using directed evolution. Biochem. Biophys. Res. Commun. 286: 126-132.
Itoh N, Kawanimi T, Liu JQ, Dairi T, Miyakoshi M, Nitta C, Kimoto Y. 2001. Cloning and biochemical characterization of Co2+-activated bromoperoxidase-esterase (perhydrolase) from Pseudomonas putida IF-3 strain. Biochim. Biophys. Acta 1545:53-66.
Jannun R, Coe EL. 1987. Bromoperoxidase from the marine snail Murex trunculis. Comp. Biochem. Physiol. 88: 917-922.
Laemmli UK. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680-685.
Meyer A, Schmid A, Held M, Westphal AH, Rothlisberger M, Kohler HP, et al. 2002. Changing the substrate reactivity of 2-hydroxybiphenyl 3-monooxygenase from Pseudomonas azelaica HBP1 by directed evolution. J. Biol. Chem. 277: 55755582.
Moore JC, Arnold FH. 1996. Directed evolution of a pnitrobenzyl esterase for aqueous-organic solvents. Nat. Biotechnol. 14: 458-467.
Murphy CD. 2003. New frontiers in biological halogenation. J. Appl. Microbiol. 94: 539-548.
Okuta A, Ohnishi K, Harayama S. 1998. PCR isolation of catechol 2,3-dioxygenase gene fragments from environmental samples and their assembly into functional genes. Gene 212:221-228.
Plat H, Kren BE, Wever R. 1987. The bromoperoxidase from the lichen Xanthoria parietina is a novel vanadium enzyme. Biochem. J. 248: 277-279.
Sheffield DJ, Smith AJ, Harry TR, Rogers LJ. 1993. Thermostability of the vanadium bromoperoxidase from Corallina officinalis. Biochem. Soc. Trans. 21: 445.
Simons BH, Barnett P, Vollenbroek EGM, Dekker HL, Muijsers AO, Messerschmidt A, Wever R. 1995. Primary structure and characterization of the vanadium chloroperoxidase from the fungus Curfularia inaequalis. Eur. J. Biochem. 229:566-574.
Suenaga H, Mitsuoka M, Ura Y, Watanabe T, Furukawa K. 2001. Directed evolution of biphenyl dioxygenase: emergence of enhanced degradation capacity for benzene, toluene, and alkylbenzenes. J. Bacteriol. 183: 5441-5444.
ten Brink HB, Dekker HL, Schoemaker HE, Wever R. 2000. Oxidation reactions catalyzed by vanadium chloroperoxidase from Curvularia inaequalis. J. Inorg. Biochem. 80: 91-98.
van Beilen JB, Duetz WA, Schmid A, Witholt B. 2003. Practical issues in the application of oxygenases. Trends Biotechnol. 21: 170-177.
Van Peé KH, Lingens F. 1985. Purification and molecular and catalytic properties of bromoperoxidase from Streptomyces phaeochromogenes. J. Gen. Microbiol. 131: 1911-1916.
Van Peé KH, Sury G, Lingens F. 1987. Purification and properties of a non-heme bromoperoxidase from Streptomyces aureofaciens. Biol. Chem. Hoppe-Seyler 368: 1225-1232.
You L, Arnold FH. 1996. Directed evolution of subtilisin E in Bacillus subtilis to enhance total activity in aqueous dimethylformamide. Protein Eng. 9: 77-83.

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Research article

J. Microbiol. Biotechnol. 2014; 24(6): 835-842

Published online June 28, 2014 https://doi.org/10.4014/jmb.1310.10070

Identification of Novel Non-Metal Haloperoxidases from the Marine Metagenome

Hui-Jeong Gwon ¹, Ide Teruhiko ², Harayama Shigeaki ² and Sang-Ho Baik ^{2, 3*}

¹Radiation Research Division for Industry & Environment, Korea Atomic Energy Research Institute, Jeongeup 580-185, Republic of Korea, ²Marine Biotechnology Institute, Heita, Kamaishi, Iwate 026-0001, Japan, ³Department of Food Science and Human Nutrition, and Research Institute of Makgeolli, Chonbuk National University, Jeonju 561-756, Republic of Korea

Received: October 21, 2013; Accepted: February 26, 2014

Abstract

Haloperoxidase (HPO, E.C.1.11.1.7) is a metal-containing enzyme oxidizing halonium species,
which can be used in the synthesis of halogenated organic compounds, for instance in the
production of antimicrobial agents, cosmetics, etc., in the presence of halides and H2O2. To
isolate and evaluate a novel non-metal HPO using a culture-independent method, a cassette
PCR library was constructed from marine seawater in Japan. We first isolated a novel HPO
gene from Pseudomonas putida ATCC11172 by PCR for constructing the chimeric HPO library
(HPO11172). HPO11172 showed each single open-reading frame of 828 base pairs coding for
276 amino acids, respectively, and showed 87% similarity with P. putida IF-3 sequences.
Approximately 600 transformants screened for chimeric genes between P. putida ATCC11173
and HPO central fragments were able to identify 113 active clones. Among them, we finally
isolated 20 novel HPO genes. Sequence analyses of the obtained 20 clones showed higher
homology genes with P. putida or Sinorhizobium or Streptomyces strains. Although the HPO A9
clone showed the lowest homology with HPO11172, clones in group B, including CS19,
showed a relatively higher homology of 80%, with 70% identy. E. coli cells expressing these
HPO chimeric genes were able to successfully bioconvert chlorodimedone with KBr or KCl as
substrate.

Keywords: haloperoxidase, marine metagenome, chimeric library, cassette PCR

Abstract

References

Almeida M, Filipe S, Humanes M, Maia MF, Melo R, Severino N, et al. 2001. Vanadium haloperoxidases from brown algae of the Laminariaceae family. Phytochemistry 57:633-642.
Iffland A, Gendreizig S, Tafelmeyer P, Johnsson K. 2001. Changing the substrate specificity of cytochrome c peroxidase using directed evolution. Biochem. Biophys. Res. Commun. 286: 126-132.
Itoh N, Kawanimi T, Liu JQ, Dairi T, Miyakoshi M, Nitta C, Kimoto Y. 2001. Cloning and biochemical characterization of Co2+-activated bromoperoxidase-esterase (perhydrolase) from Pseudomonas putida IF-3 strain. Biochim. Biophys. Acta 1545:53-66.
Jannun R, Coe EL. 1987. Bromoperoxidase from the marine snail Murex trunculis. Comp. Biochem. Physiol. 88: 917-922.
Laemmli UK. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680-685.
Meyer A, Schmid A, Held M, Westphal AH, Rothlisberger M, Kohler HP, et al. 2002. Changing the substrate reactivity of 2-hydroxybiphenyl 3-monooxygenase from Pseudomonas azelaica HBP1 by directed evolution. J. Biol. Chem. 277: 55755582.
Moore JC, Arnold FH. 1996. Directed evolution of a pnitrobenzyl esterase for aqueous-organic solvents. Nat. Biotechnol. 14: 458-467.
Murphy CD. 2003. New frontiers in biological halogenation. J. Appl. Microbiol. 94: 539-548.
Okuta A, Ohnishi K, Harayama S. 1998. PCR isolation of catechol 2,3-dioxygenase gene fragments from environmental samples and their assembly into functional genes. Gene 212:221-228.
Plat H, Kren BE, Wever R. 1987. The bromoperoxidase from the lichen Xanthoria parietina is a novel vanadium enzyme. Biochem. J. 248: 277-279.
Sheffield DJ, Smith AJ, Harry TR, Rogers LJ. 1993. Thermostability of the vanadium bromoperoxidase from Corallina officinalis. Biochem. Soc. Trans. 21: 445.
Simons BH, Barnett P, Vollenbroek EGM, Dekker HL, Muijsers AO, Messerschmidt A, Wever R. 1995. Primary structure and characterization of the vanadium chloroperoxidase from the fungus Curfularia inaequalis. Eur. J. Biochem. 229:566-574.
Suenaga H, Mitsuoka M, Ura Y, Watanabe T, Furukawa K. 2001. Directed evolution of biphenyl dioxygenase: emergence of enhanced degradation capacity for benzene, toluene, and alkylbenzenes. J. Bacteriol. 183: 5441-5444.
ten Brink HB, Dekker HL, Schoemaker HE, Wever R. 2000. Oxidation reactions catalyzed by vanadium chloroperoxidase from Curvularia inaequalis. J. Inorg. Biochem. 80: 91-98.
van Beilen JB, Duetz WA, Schmid A, Witholt B. 2003. Practical issues in the application of oxygenases. Trends Biotechnol. 21: 170-177.
Van Peé KH, Lingens F. 1985. Purification and molecular and catalytic properties of bromoperoxidase from Streptomyces phaeochromogenes. J. Gen. Microbiol. 131: 1911-1916.
Van Peé KH, Sury G, Lingens F. 1987. Purification and properties of a non-heme bromoperoxidase from Streptomyces aureofaciens. Biol. Chem. Hoppe-Seyler 368: 1225-1232.
You L, Arnold FH. 1996. Directed evolution of subtilisin E in Bacillus subtilis to enhance total activity in aqueous dimethylformamide. Protein Eng. 9: 77-83.

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Research article

Identification of Novel Non-Metal Haloperoxidases from the Marine Metagenome

Abstract

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Research article

Identification of Novel Non-Metal Haloperoxidases from the Marine Metagenome

Abstract

References