A Novel Phenanthrene Dioxygenase from Nocardioides sp. Strain KP7: Expression in Escherichia coli

Journal of Bacteriology - Tập 182 Số 8 - Trang 2134-2141 - 2000
Atsushi Saito1, Tokuro Iwabuchi1, Shigeaki Harayama1
1Marine Biotechnology Institute, Kamaishi Laboratories, Kamaishi, Iwate 026-0001, Japan

Tóm tắt

ABSTRACT Nocardioides sp. strain KP7 grows on phenanthrene but not on naphthalene. This organism degrades phenanthrene via 1-hydroxy-2-naphthoate, o -phthalate, and protocatechuate. The genes responsible for the degradation of phenanthrene to o -phthalate ( phd ) were found by Southern hybridization to reside on the chromosome. A 10.6-kb DNA fragment containing eight phd genes was cloned and sequenced. The phdA , phdB , phdC , and phdD genes, which encode the α and β subunits of the oxygenase component, a ferredoxin, and a ferredoxin reductase, respectively, of phenanthrene dioxygenase were identified. The gene cluster, phdAB , was located 8.3 kb downstream of the previously characterized phdK gene, which encodes 2-carboxybenzaldehyde dehydrogenase. The phdCD gene cluster was located 2.9 kb downstream of the phdB gene. PhdA and PhdB exhibited moderate (less than 60%) sequence identity to the α and β subunits of other ring-hydroxylating dioxygenases. The PhdC sequence showed features of a [3Fe-4S] or [4Fe-4S] type of ferredoxin, not of the [2Fe-2S] type of ferredoxin that has been found in most of the reported ring-hydroxylating dioxygenases. PhdD also showed moderate (less than 40%) sequence identity to known reductases. The phdABCD genes were expressed poorly in Escherichia coli , even when placed under the control of strong promoters. The introduction of a Shine-Dalgarno sequence upstream of each initiation codon of the phdABCD genes improved their expression in E. coli. E. coli cells carrying phdBCD or phdACD exhibited no phenanthrene-degrading activity, and those carrying phdABD or phdABC exhibited phenanthrene-degrading activity which was significantly less than that in cells carrying the phdABCD genes. It was thus concluded that all of the phdABCD genes are necessary for the efficient expression of phenanthrene-degrading activity. The genetic organization of the phd genes, the phylogenetically diverged positions of these genes, and an unusual type of ferredoxin component suggest phenanthrene dioxygenase in Nocardioides sp. strain KP7 to be a new class of aromatic ring-hydroxylating dioxygenases.

Từ khóa


Tài liệu tham khảo

10.1093/nar/25.17.3389

10.1128/JB.180.15.3954-3966.1998

Ausubel F. M. Brent R. Kingston R. E. Moore D. D. Seidman J. G. Smith J. A. Struhl K. Current protocols in molecular biology Suppl. 11. unit 16.2 1990 John Wiley & Sons New York N.Y

Batie C. J. Ballou D. P. Correll C. C. Phthalate dioxygenase reductase and related flavin-iron-sulfur containing electron transferases Chemistry and biochemistry of flavoenzymes Muller F. 3 1992 543 556 CRC Press Boca Raton, Fla

Bergeron J. Ahmad D. Barriault D. Larose A. Sylvestre M. Powlowski J. Identification and mapping of the gene translation products involved in the first steps of the Comamonas testosteroni B-356 biphenyl/chlorobiphenyl biodegradation pathway.Can. J. Microbiol.401994743753

10.1016/S0378-1119(99)00241-3

Brosius J. Superpolylinkers in cloning and expression vectors.DNA81989759777

Cerniglia C. E. Heitkamp M. A. Microbial degradation of polycyclic aromatic hydrocarbons (PAH) in the aquatic environment Metabolism of polycyclic aromatic hydrocarbons in the aquatic environment. Varanasi U. 1989 41 68 CRC Press Inc. Boca Raton Fla

10.1128/jb.176.9.2492-2501.1994

10.1128/jb.175.21.6890-6901.1993

10.1128/jb.176.24.7757-7762.1994

10.1128/jb.178.5.1351-1362.1996

10.1128/jb.174.23.7542-7554.1992

Elliott J. I. Yang S. S. Ljungdahl L. G. Travis J. Reilly C. F. Complete amino acid sequence of the 4Fe-4S, thermostable ferredoxin from Clostridium thermoaceticum.Biochemistry21198232943298

10.1128/jb.174.9.2903-2912.1992

Felsenstein J. Phylogeny Inference Package version 3.5c. 1993 Department of Genetics University of Washington Seattle Wash (Distributed by the author.)

10.1038/387394a0

10.1128/JB.180.9.2522-2530.1998

Gibson D. T. Subramanian V. Microbial degradation of aromatic hydrocarbons Microbial degradation of organic compounds. Gibson D. T. 1984 181 252 Marcel Dekker Inc. New York N.Y

Harayama S. Polycyclic aromatic hydrocarbon bioremediation design.Curr. Opin. Biotechnol.81997268273

Harayama S. Kok M. Neidle E. L. Functional and evolutionary relationships among diverse oxygenases.Annu. Rev. Microbiol.461992565601

10.1128/jb.176.15.4790-4793.1994

10.1128/jb.169.11.5174-5179.1987

10.1128/jb.179.20.6488-6494.1997

10.1128/JB.180.4.945-949.1998

Iwabuchi T. Harayama S. Biochemical and molecular characterization of 1-hydroxy-2-naphthoate dioxygenase from Nocardioides sp. KP7.J. Biol. Chem.273199883328336

Iwabuchi T. Inomata-Yamauchi Y. Katsuta A. Harayama S. Isolation and characterization of marine Nocardioides capable of growing and degrading phenanthrene at 42°C.J. Mar. Biotechnol.619988690

Kauppi B. Lee K. Carredano E. Parales R. E. Gibson D. T. Eklund H. Ramaswamy S. Structure of an aromatic-ring-hydroxylating dioxygenase-naphthalene 1,2-dioxygenase.Structure61998571586

10.1128/aem.43.2.458-461.1982

10.1128/jb.176.8.2439-2443.1994

Kurkela S. Lehvaslaiho H. Palva E. T. Teeri T. H. Cloning, nucleotide sequence and characterization of genes encoding naphthalene dioxygenase of Pseudomonas putida strain NCIB9816.Gene731988355362

10.1128/JB.181.2.531-540.1999

10.1128/JB.181.9.2675-2682.1999

10.1128/aem.61.6.2079-2085.1995

Mason J. R. Cammack R. The electron-transport proteins of hydroxylating bacterial dioxygenases.Annu. Rev. Microbiol.461992277305

Nelson K. E. Clayton R. A. Gill S. R. Gwinn M. L. Dodson R. J. Haft D. H. Hickey E. K. Peterson J. D. Nelson W. C. Ketchum K. A. McDonald L. Utterback T. R. Malek J. A. Linher K. D. Garrett M. M. Stewart A. M. Cotton M. D. Pratt M. S. Phillips C. A. Richardson D. Heidelberg J. Sutton G. G. Fleischmann R. D. White O. Salzberg S. L. Smith H. O. Venter J. C. Fraser C. M. Evidence for lateral gene transfer between Archaea and bacteria from genome sequence of Thermotoga maritima.Nature3991999323329

O'Keefe D. P. Gibson K. J. Emptage M. H. Lenstra R. Romesser J. A. Litle P. J. Omer C. A. Ferredoxins from two sulfonylurea herbicide monooxygenase systems in Streptomyces griseolus.Biochemistry301991447455

Page R. D. TreeView: an application to display phylogenetic trees on personal computers.Comput. Appl. Biosci.121996357358

Saito A. Iwabuchi T. Harayama S. Characterization of genes for enzymes involved in the phenanthrene degradation in Nocardioides sp. KP7.Chemosphere38199913311337

Sambrook J. Fritsch E. F. Maniatis T. Molecular cloning: a laboratory manual 2nd ed. 1989 Cold Spring Harbor Laboratory Cold Spring Harbor N.Y

Shepherd J. M. Lloyd-Jones G. Novel carbazole degradation genes of Sphingomonas CB3: sequence analysis, transcription, and molecular ecology.Biochem. Biophys. Res. Commun.2471998129135

Simon M. J. Osslund T. D. Saunders R. Ensley B. D. Suggs S. Harcourt A. Suen W. C. Cruden D. L. Gibson D. T. Zylstra G. J. Sequences of genes encoding naphthalene dioxygenase in Pseudomonas putida strains G7 and NCIB 9816-4.Gene12719933137

Simons R. Priefer U. Puhler A. A broad host range mobilization system of in vivo genetic engineering: transposon mutagenesis in gram negative bacteria.Bio/Technology11983784790

10.1128/JB.169.12.5789-5794.1987

10.1016/0076-6879(90)85008-C

Sylvestre M. Sirois M. Hurtubise Y. Bergeron J. Ahmad D. Shareck F. Barriault D. Guillemette I. Juteau J. M. Sequencing of Comamonas testosteroni strain B-356-biphenyl/chlorobiphenyl dioxygenase genes: evolutionary relationships among Gram-negative bacterial biphenyl dioxygenases.Gene1741996195202

10.1016/S0021-9258(18)42908-0

10.1128/jb.176.8.2444-2449.1994

Thompson J. D. Higgins D. G. Gibson T. J. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice.Nucleic Acids Res.11199446734680

Treadway S. L. Yanagimachi K. S. Lankenau E. Lessard P. A. Stephanopoulos G. Sinskey A. J. Isolation and characterization of indene bioconversion genes from Rhodococcus strain I24.Appl. Microbiol. Biotechnol.511999786793

Trower M. K. Lenstra R. Omer C. Buchholz S. E. Sariaslani F. S. Cloning, nucleotide sequence determination and expression of the genes encoding cytochrome P-450soy (soyC) and ferredoxinsoy (soyB) from Streptomyces griseus.Mol. Microbiol.6199221252134

Wang P. L. Donaire A. Zhou Z. H. Adams M. W. La Mar G. N. Molecular model of the solution structure for the paramagnetic four-iron ferredoxin from the hyperthermophilic archaeon Thermococcus litoralis.Biochemistry3519961131911328

Wierenga R. K. Terpstra P. Hol W. G. J. Prediction of the occurrence of the ADP-binding bab-fold in proteins, using an amino acid sequence fingerprint.J. Mol. Biol.1871986101107

10.1128/jb.176.8.2158-2164.1994

10.1016/0378-1119(85)90120-9

Zhou Z. H. Adams M. W. W. Proton NMR investigation of the oxidized three-iron clusters in the ferredoxins from the hyperthermophilic archae Pyrococcus furiosus and Thermococcus litoralis.Biochemistry3119921195211962

Zhou Z. H. Adams M. W. W. Site-directed mutations of the 4Fe-ferredoxin from the hyperthermophilic archaeon Pyrococcus furiosus: role of the cluster-coordinating aspartate in physiological electron transfer reactions.Biochemistry3619971089210900

Zylstra G. J. Gibson D. T. Toluene degradation by Pseudomonas putida F1. Nucleotide sequence of the todC1C2BADE genes and their expression in Escherichia coli.J. Biol. Chem.26419891494014946

Thông tin
Thông tin xuất bản

Journal of Bacteriology

Tập 182 Số 8

2134-2141

Thông tin tác giả