| 摘要(英) |
Alkylphenol polyethoxylate (APEOn), including octylphenol polyethoxylate (OPEOn) and nonylphenol polyethoxylate (NPEOn), are typical polyethylene glycol (PEG) type non-ionic surfactants. The structures of APEOn are a hydrophilic ethylene oxide (EO) chain attached to a hydrophobic alkylphenol at para-position. They are very important surfactants with industrial, agricultural, and domestic applications. The majority of environmental APEOn comes from wastewater treatment, plant discharge, pesticides and herbicides on farms. When APEOn are released into environment, the EO chains of APEOn are cleaved to APEOn (n≦3), APECn (n≦3) and alkylphenol (AP) which are known to cause estrogenic effects on aquatic organisms, birds, and mammals as environmental hormoens. Pseudomonas nitroreducens TX1, isolated in Taiwan, is shown to be able to grow on 0.05~20％(v/v) of OPEOn as sole carbon source. Liquid chromatography-mass spectrometry analysis showed that the biodegradation mechanism of OPEOn and dodecyl octaethoxylate (AEO8) by strain TX1 was through a sequential cleavage of the EO chain and accompany with the formation of carboxylated OPEOn and AEO8. Previously, P. nitroreducens TX1 was grown on minimal salts basal (MSB) medium containing 0.5％ OPEOn as sole carbon source and analyzed by a subtractive proteome comparisons against a succinate-grown proteome. An alcohol dehydrogenase containing pyrroloquinoline quinine (PQQ) and aldehyde dehydrogenase (ALDH) were shown to be up-regulated for 15-and 10-folds, respectively. Moreover, an alcohol dehydrogenase (PQQ) was shown to degrade PEG proposed by other group. Therefore, we cloned ADH(PQQ) and ALDH for expression in Escherichia coli. However, the ADH expressed in E. coli is mostly in the membrane fraction and we can not detect the activity in this fraction. We then identified an OPEOn degrading enzyme (ADH) from strain TX1 and found that ADH react on OPEOn is neither NAD+ nor PQQ-dependent enzymes. We can not determine the cofactor of this enzyme and temporarily named ADH. The alcohol dehydrogenase activity was up-regulated for 3 folds in cell grown on 0.5% OPEOn compared to that on 0.5% succinate, indicating OPEOn induced a higher expression of ADH. The activities of ADH in crude extract and in membrane are 74 and 26 percents, respectively. ADH in crude extract, has activity to AEO8, OPEOn and NPEOn (9~11 mU/mg of crude extract) but has no activity to PEG. Therefore, the ADH from P. nitroreducens TX1 is different from PEG dehydrogenase from Sphingomonas. terrae published previously. In addition, ADH has no activity to methanol, ethanol, butanol, benzyl alcohol, but the ADH also is a dehydrogenase for nonyl-aldehyde (22 mU/mg), but not on formaldehyde and benzaldehyde, indicating its role in the formation of APEC. The ADH from strain TX1 is specifically to catalyze OPEOn to form OPECn. By ionic exchange chromatography, the pI of ADH is estimated to be around 5. The first purification step is by anionic exchange chromatography. The recovery is 22% from the first step. Second step is by gel filtration and the recovery is 3%. Chromatofocusing was used as the third step for ADH purification. The pI of ADH is 5.8. Substrates specificity of the partial purified ADH from second-step purification is the same as ADH in crude extract. In the future work, the ADH will be cloned to analyze its products analyzed by LC-MS to confirm the role of ADH in the OPEOn degradation pathway in P. nitroreducens TX1. |
| 參考文獻 |
丁望賢、吳建誼、周瓊瑤、王正雄。2000。壬基苯酚及其相關化學物質在台灣水環境中之分析與調查，第一屆環境荷爾蒙研討會論文集。p.153-5。
邱凡峰。2005。以功能性蛋白質體學研究Pseudomonas nitroreducens TX1生長於辛基苯酚聚氧乙基醇之代謝與逆境反應。國立中央大學生命科學研究所碩士論文。
李美慧。2000。常見環境荷爾蒙物質及其影響。第一屆環境荷爾蒙研討會論文集。p.4-14。
洪國展。2004。分解辛基苯酚聚氧乙基醇之耗氧酵素（二氫硫辛醯胺脫氫酶）的純化與定性。國立中央大學生命科學研究所碩士論文。
陳錫金。2005。界面活性劑Octylphenol Polyethoxylates生物降解與復育之研究。國立中央大學環境工程研究所博士論文。
廖明隆 譯。1994。界面化學與界面活性劑。文原書局。p.13-35。
謝孝正。2004。Pseudomonas putida TX2分解辛基苯酚聚氧乙基醇其具雌激素活性代謝物之研究。國立中央大學生命科學研究所碩士論文。
Ahel, M., F. E. Scully, J. Hoigne and W. Giger. 1994. Photochemical degradation of nonylphenol and nonylphenolpolyethoxylatesin natural-water. Chemosphere. 28:1361-68.
Bradford, M. M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 72:248-54.
Brand, N., G. Mailhot, and M. Bolte. 1998. Degradation photoinduced by Fe(III): method of alkylphenol ethoxylates removal in Water. Environ. Sci. Technol. 32:2715-20.
Chen, H. J., S. L. Huang, and D. H. Tseng. 2004. Aerobic biotransformation of octylphenol polyethoxylate surfactant in soil microcosms. Environ. Technol. 25:201-10.
Hauthal, H. G. 1992. Trends in surfactants. Chim Oggi. 10: 9-13.
Herold DA, Keil K, Bruns DE. 1989. Oxidation of polyethylene glycols by alcohol dehydrogenase. Biochem Pharmacol. 38:73-76.
Hoffmeister M, Piotrowski M, Nowitzki U, Martin W. 2005. Mitochondrial trans-2-enoyl-CoA reductase of wax ester fermentation from Euglena gracilis defines a new family of enzymes involved in lipid synthesis. J. Biol. Chem. 280(6):4329-38.
Huang, S. L., C. J. Yang, G. L. Guo, and S. H. Chou. 2004. Isolation, identification and properties of bacterial strains degrading octylphenol polyethoxylates. Taiwan Journal of Agricultural Chemistry and Food Science. 42:356-365.
John, D. M., White. G. F. 1998. Mechanism for biotransformation of nonylphenol polyethoxylates to xenoesterogens in Pseudomonas putida. J. Bacteriol. 180:4332-8.
Jon GA de, Geerlof HA, Stoorvogel J, Jongejan JA, Vries A de, Duine JA. 1995. Quinohaemoprotein ethanol dehydrogenase from Comamonas testosteroni: purification, characterization, and reconstitution of the apoenzyme. Eur J Biochem. 230:899-905.
Kawai, F., T. Kimura, Y. Tani, H. Yamada, T. Ueno, and H, Fukami. 1983. Identification of reaction products of polyethylene glycol dehydrogenase. Agric. Biol. Chem. 47:1669-1671.
Kawai, F. 2002. Microbial degradation of polyethers. Appl Microbiol Biotechnol. 58:30-8.
Kawai, F., T. Kimura, Y. Tani, H. Yamada, and M. Kurachi. 1980. Purification and characterization of polyethylene glycol dehydrogenase involved in the bacterial metabolism of polyethylene glycol. Appl Environ Microbiol. 40:701-5.
Kerem, Z., hammel, and K. E. Hammel. 1999. Biodegradative mechanism of the brown rot basidiomycete Gloeophyllum trabeum: evidence for an extracellular hydroquinone-driven fenton reaction. FEBS Lett. 446:49-54.
Liu, X., Tani, A., Kimbara, K. 2007. Xenoestrogenic short ethoxy chain nonylphenol is oxidized by a flavoprotein alcohol dehydrogenase from Ensifer sp. strain AS08. Appl Microbiol Biotechnol. 73:1414-1422.
Montgomery-Brown, J., M. Reinhard. 2003. Occurrence and behavior of alkylphenol polyethoxylates in the environment. Enviro. Eng. Sci. 20:471-86.
Naylor., C. G. 1995. Environmental fate and safety of nonylphenol ethoxylates. Textile Chemist. Colorist. 27:29-33.
Ohta, T., Tani, A. 2005. A novel nicotinoprotein aldehyde dehydrogenase involved in polyethylene glycol degradation. Appl Microbiol Biotechnol. 68:639-646.
Reichmann, P., and H. Gorisch. 1993. Cytochrome c550 from Pseudomonas aeruginosa. Biochem J. 289(Pt 1):173-8.
Reid, MF., and Fewson, CA. 1994. Molecular characterization of microbial alcohol dehydrogenase. Crit Rev Microbiol. 20(1):13-56.
Rupp, M., and H. Gorisch. 1988. Purification, crystallisation and characterization of quinoprotein ethanol dehydrogenase from Pseudomonas aeruginosa. Biol. Chem. Hoppe. Seyler. 369:431-9.
Sugimoto M, Tanabe M, Hataya M, Enokibara S, Duine JA, Kawai F. 2001. The first step in polyethylene glycol degradation by sphingomonads proceeds via a flavoprotein (FAD-containing)alcohol dehydrogenase. J Bacteriol. 183:6694-6698.
Tachibana, S., F. Kawai, and M. Yasuda. 2002. Heterogeneity of dehydrogenases of Stenotrophomonas maltophilia showing dye-linked activity with polypropylene glycols. Biosci. Biotechnol. Biochem. 66:737-42.
Tanenbaum, D. M., Y. Wang, S. Williams and P. Sigler. 1998. Crystallographic comparison of the estrogen and progesterone receptor’s ligand binding domains. Proc. Natl. Acad. Sci. 95:5998-6003.
Tasaki, Y., Yoshikawa, H., and Tamura, H. 2006. Isolation and characterization of alcohol dehydrogenase gene from the octylphenol polyethoxylate degrader Pseudomonas putida S-5. Biosci. Biotechnol. Biochem. 70(8):1855-1863.
Tracy E, Stites., Tracey R, Sih., Robert B, Rucker. 2000. Synthesis of [14C]pyrroloquinoline quinone (PQQ) in E. coli using genes for PQQ synthesis from K. pneumoniae. Biochimica et Biophysica Acta. 1524:247-252.
Van Ginkel, C. G. 1996. Complete degradation of xenobiotic surfactants by consortia of aerobic microorganisms. Biodegradation. 7:151-64.
Von Tigerstrom RG., Razzell WE. 1968. Aldehyde dehydrogenase. I. Purification and properties of the enzyme from Pseudomonas aeruginosa. J Biol Chem. 243(10):2691-2702.
Yamashita, M., Tani, A., Kawai, F. 2005. A new ether bond-splitting enzyme found in Gram-positive polyethylene glycol 6000-utilizing bacterium, Pseudonocardia sp.strain K1. Appl Microbiol Biotechnol. 66:174-179.
Yasuda M, Cherepanov A, Duine JA. 1996. Polyethylene glycol dehydrogenase activity of Rhodopseudomonas acidophila derives from a type I quinohaemoprotein alcohol dehydrogenase. FEMS Microbiol Lett. 138:23-38.
Zarnt G, Schraeder T, Andreesen JR. 1997. Degradation of tetrahydrofurfuryl alcohol by Ralstonia eutropha is initiated by an Inducible pyrroloquinoline quinone-dependent alcohol dehydrogenase. Appl Environ Microbiol. 63:4891-4898. |
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