Skip to main content

Advertisement

Log in

Biodegradation of dioxins by recombinant Escherichia coli expressing rat CYP1A1 or its mutant

  • Environmental Biotechnology
  • Published:
Applied Microbiology and Biotechnology Aims and scope Submit manuscript

Abstract

Among polychlorinated dibenzo-p-dioxins (PCDDs), 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TetraCDD) is the most toxic one. Recently, we reported that rat CYP1A1 mutant, F240A, expressed in yeast showed metabolic activity toward 2,3,7,8-TetraCDD. In this study, we successfully expressed N-terminal truncated P450s (Δ1A1 and ΔF240A) in Escherichia coli cells. Kinetic analysis using membrane fractions prepared from the recombinant E. coli cells revealed that ΔF240A has enzymatic properties similar to F240A expressed in yeast. The metabolism of PCDDs by recombinant E. coli cells expressing both ΔF240A and human NADPH-P450 reductase was also examined. When 2,3,7-TriCDD was added to the E.coli cell culture at a final concentration of 10 μM, approximately 90% of the 2,3,7-TriCDD was converted into multiple metabolites within 8 h. These results indicate the possible application of prokaryotic cells expressing ΔF240A to the bioremediation of PCDD-contaminated soil.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Subscribe and save

Springer+
from $39.99 /Month
  • Starting from 10 chapters or articles per month
  • Access and download chapters and articles from more than 300k books and 2,500 journals
  • Cancel anytime
View plans

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

Explore related subjects

Discover the latest articles and news from researchers in related subjects, suggested using machine learning.

References

  • Aoyama Y, Yoshida Y, Kubota S, Kumaoka H, Furumichi A (1978) NADPH-cytochrome P-450 reductase of yeast microsomes. Arch Biochem Biophys 185:362–369

    Article  CAS  PubMed  Google Scholar 

  • Armengaud J, Happe B, Timmis K (1998) Genetic analysis of dioxin dioxygenase of Sphingomonas sp. strain RW1: catabolic genes dispersed on the genome. J Bacteriol 180:3954–3966

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Barnes HJ, Arlotto MP, Waterman M (1991) Expression and enzymatic activity of recombinant cytochrome P450 17α-hydroxylation in Escherichia coli. Proc Natl Acad Sci U S A 88:5597–5601

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bumpus JA, Tien M, Wright D, Aust SD (1985) Oxidation of persistent environmental pollutants by a white rot fungus. Science 228:1434–1436

    Article  CAS  PubMed  Google Scholar 

  • Bunge M, Adrian L, Kraus A, Opel M, Lorenz WG, Andreesen JR, Görisch H, Lechner U (2003) Reductive dehalogenation of chlorinated dioxins by an anaerobic bacterium. Nature 421:357–360

    Article  CAS  PubMed  Google Scholar 

  • Cosme J, Johnson EF (2000) Engineering microsomal cytochrome P450 2C5 to be a soluble, monomeric enzyme. Mutations that alter aggregation, phospholipid dependence of catalysis, and membrane binding. J Biol Chem 275:2545–2553

    Article  CAS  PubMed  Google Scholar 

  • Graham-Lorence S, Amarneh B, White RE (1995) A three-dimensional model of aromatase cytochrome P450. Protein Sci 4:1065–1080

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Guo Z, Gilliam EMJ, Ohmori S, Tukey RH, Guengerich FP (1994) Expression of modified human cytochrome P450 1A1 in Escherichia coli: effects of 5’ substitution, stabilization, purification, spectral characterization, and catalytic properties. Arch Biochem Biophys 312:436–446

    Article  CAS  PubMed  Google Scholar 

  • Enguita FJ, de la Fuente JL, Martin JF, Liras P (1996) An inducible expression system of histidine-tagged proteins in Streptomyces lividans for one-step purification by Ni2+ affinity chromatography. FEMS Microbiol Lett 137:151–156

    Article  Google Scholar 

  • Fornwald JA, Donovan MJ, Geber R, Keller J, Taylor DP, Arcuri EJ, Brawner ME (1993) Soluble forms of the human T cell receptor CD4 are efficiently expressed by Streptomyces lividans. Biotechnology 11:1031–1036

    CAS  PubMed  Google Scholar 

  • Inouye K, Shinkyo R, Takita T, Ohta M, Sakaki T (2002) Metabolism of polychlorinated dibenzo-p-dioxins (PCDDs) by human cytochrome P450-dependent monooxygenase systems. J Agric Food Chem 50:5496–5502

    Article  CAS  PubMed  Google Scholar 

  • Iwata H, Fujita K, Kushida H, Suzuki A, Konno Y, Nakamura K, Fujino A, Kamataki T (1998) High catalytic activity of human cytochrome P450 co-expressed with human NADPH-cytochrome P450 reductase in Escherichia coli. Biochem Pharmacol 55:1315–1325

    Article  CAS  PubMed  Google Scholar 

  • Jenkins CM, Waterman MR (1994) Flavodoxin and NADPH-flavodoxin reductase from Escherichia coli support bovine cytochrome P450c17 hydroxylase activities. J Biol Chem 269:27401–27408

    CAS  PubMed  Google Scholar 

  • Kimura N, Urushigawa Y (2001) Metabolism of dibenzo-p-dioxin and chlorinated dibenzo-p-dioxin by a gram-positive bacterium, Rhodococcus opacus SAO101. J Biosci Bioeng 92:138–143

    Article  CAS  PubMed  Google Scholar 

  • Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the folin phenol reagent. J Biol Chem 193:265–275

    CAS  PubMed  Google Scholar 

  • Mast N, Andersson U, Nakayama K, Bijorkhem I, Pikuleva A (2004) Expression of human cytochrome P450 46A1 in Escherichia coli: effects of N- and C-terminal modifications. Arch Biochem Biophys 428:99–108

    Article  CAS  PubMed  Google Scholar 

  • Murtazina D, Puchkaev AV, Schein CH, Oezguen N, Braun W, Nanavati A, Pikuleva IA (2002) Membrane–protein interactions contribute to efficient 27-hydroxylation of cholesterol by mitochondrial cytochrome P450 27A1. J Biol Chem 277:37582–37589

    Article  CAS  PubMed  Google Scholar 

  • Nakashima N, Tamura T (2004a) A novel system of expressing recombinant proteins over a wide temperature range from 4 to 35 degrees C. Biotechnol Bioeng 86:136–148

    Article  CAS  PubMed  Google Scholar 

  • Nakashima N, Tamura T (2004b) Isolation and characterization of a rolling-circle-type plasmids from Rhodococcus erythropolis and application of the plasmid to multiple-recombinant protein expression. Appl Environ Microbiol 70:1181–1189

    Article  Google Scholar 

  • Nojiri H, Nam JW, Kosaka M, Morii K, Takemura IT, Furihata K, Yamane H, Omori T (1999) Diverse oxygenations catalyzed by carbazole 1,9a-dioxygenase from Pseudomonas sp. Strain CA10. J Bacteriol 181:3105–3113

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ohta Y, Sakaki T, Yabusaki Y, Ohkawa H, Kawato S (1994) Rotation and membrane topology of genetically expressed methylcolanthrene-inducible cytochrome P-4501A1 lacking the N-terminal hydrophobic segment in yeast microsomes. J Biol Chem 269:15597–15600

    CAS  PubMed  Google Scholar 

  • Olins PO, Rangwala SH (1989) A novel sequence element derived from Bacteriophage T7 mRNA acts as an enhancer of translation of the lacZ gene in Escherichia coli. J Biol Chem 264:16973–1697

    Google Scholar 

  • Omura T, Sato R (1964) The carbon monoxide-binding pigment of liver microsomes: II. Solubilization, purification, and properties. J Biol Chem 239:2379–238

    CAS  PubMed  Google Scholar 

  • Orihara K, Yamazaki T, Shinkyo R, Sakaki T, Inouye K, Tsukamoto A, Sugiura J, Shishido K (2005) Rat cytochrome P450-mediated transformation of dichlorodibenzo-p-dioxins by recombinant white-rot basidiomycete Coriolus hirsutus. Appl Microbiol Biotechnol 69(1):22–28

    Article  CAS  PubMed  Google Scholar 

  • Parikh A, Guengerich FP (1997) Drug metabolism by Escherichia coli expressing human cytochromes P450. Nat Biotechnol 15:784–788

    Article  CAS  PubMed  Google Scholar 

  • Sato SI, Nam JW, Kasuga K, Nojiri H, Yamane H, Omori T (1997) Identification and characterization of genes encoding carbazol 1,9a-dioxygenase in Pseudomonas sp. Strain CA10. J Bacteriol 179:4850–4858

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sakaki T, Soga A, Yabusaki Y, Ohkawa H (1984) Characterization of three forms of cytochrome P-450 isolated from liver microsomes of rats treated with 3-methylcholamthrene. J Biochem 96:117–126

    Article  CAS  PubMed  Google Scholar 

  • Sakaki T, Kominami S, Takemori S, Ohkawa H, Akiyoshi-Shibata M, Yabusaki Y (1994) Kinetic studies on a genetically engineered fused enzyme between rat cytochrome P4501A1 and yeast NADPH-P450 reductase. Biochemistry 33:4933–4939

    Article  CAS  PubMed  Google Scholar 

  • Sakaki T, Shinkyo R, Takita T, Ohta M, Inouye K (2002) Biodegradation of polychlorinated dibenzo-p-dioxins by recombinant yeast expressing rat CYP1A subfamily. Arch Biochem Biophys 401:91–98

    Article  CAS  PubMed  Google Scholar 

  • Shinkyo R, Sakaki T, Ohta M, Inouye K (2003a) Metabolic pathways of dioxin by CYP1A1: species difference between rat and human CYP1A subfamily in the metabolism of dioxins. Arch Biochem Biophys 409:180–187

    Article  CAS  PubMed  Google Scholar 

  • Shinkyo R, Sakaki T, Takita T, Ohta M, Inouye K (2003b) Generation of 2,3,7,8-TCDD-metabolizing enzyme by modifying rat CYP1A1 through site-directed mutagenesis. Biochem Biophys Res Commun 308:511–517

    Article  CAS  PubMed  Google Scholar 

  • Spratt JM, Britton WJ, Triccas JA (2003) Identification of strong promoter elements of Mycobacterium smegmatis and their utility for foreign gene expression in mycobacteria. FEMS Microbiol Lett 224:139–142

    Article  CAS  PubMed  Google Scholar 

  • Sulistyaningdyah WT, Ogawa J, Li QS, Shinkyo R, Sakaki T, Inouye K, Schmid RD, Shimizu S (2004) Metabolism of polychlorinated dibenzo-p-dioxins by cytochrome P450 BM-3 and its mutant. Biotechnol Lett 26:1857–1860

    Article  CAS  PubMed  Google Scholar 

  • Vail RB, Homann MJ, Hanna I, Zaks A (2005) Preparative synthesis of drug metabolites using human cytochrome P450s 3A4, 2C9, and 1A2 with NADPH-P450 reductase expressed in Escherichia coli. J Ind Microbiol Biotechnol 32:67–74

    Article  CAS  PubMed  Google Scholar 

  • Williams PA, Comse J, Sridhar V, Johnson EF, McRee DE (2000) Mammalian microsomal cytochrome P450 monooxygenase: structural adaptations for membrane binding and functional diversity. Mol Cell 5:121–131

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Toshiyuki Sakaki.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Shinkyo, R., Kamakura, M., Ikushiro, Si. et al. Biodegradation of dioxins by recombinant Escherichia coli expressing rat CYP1A1 or its mutant. Appl Microbiol Biotechnol 72, 584–590 (2006). https://doi.org/10.1007/s00253-005-0286-1

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00253-005-0286-1

Keywords