Purification and biochemical characterization of manganesecontaining superoxide dismutase from deep-sea thermophile <i>Geobacillus</i> sp. EPT3

ZHU Yanbing; LI Hebin; NI Hui; LIU Jingwen; XIAO Anfeng; CAI Huinong

doi:10.1007/s13131-014-0534-2

深海嗜热菌Geobacillus sp. EPT3锰超氧化物歧化酶的纯化与生物化学鉴定

doi: 10.1007/s13131-014-0534-2

1.
集美大学食品与生物工程学院, 集美大学, 厦门 361021, 中国;福建省食品微生物与酶工程重点实验室, 厦门 361021, 中国;福建省高校食品微生物与酶工程技术研究中心, 厦门 361012, 中国
2.
厦门医学高等专科学校, 厦门 361008, 中国
3.
集美大学食品与生物工程学院, 集美大学, 厦门 361021, 中国

基金项目: The National Natural Science Foundation of China under contract No. 31371751; the Science and Technology Program of Xiamen, China under contract No. 201303120001; the Foundation for Innovative Research Team of Jimei University, China under contract No. 2010A006.

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- 收稿日期: 2013-03-15
- 修回日期: 2013-09-13

Purification and biochemical characterization of manganesecontaining superoxide dismutase from deep-sea thermophile Geobacillus sp. EPT3

1.
College of Food and Biological Engineering, Jimei University, Xiamen 361021, China;Fujian Provincial Key Laboratory of Food Microbiology and Enzyme Engineering, Xiamen 361021, China;Research Center of Food Microbiology and Enzyme Engineering Technology of Fujian Province, Xiamen 361021, China
2.
Xiamen Medical College, Xiamen 361008, China
3.
College of Food and Biological Engineering, Jimei University, Xiamen 361021, China

摘要

摘要: 热稳定SOD是一种在生物技术应用中有潜力的酶.本研究中,嗜热菌Geobacillus sp. EPT3分离自东太平洋一个深海热液区.经硫酸铵分级沉淀、DEAE-Sepharose层析及Phenyl-Sepharose层析,从该菌株中纯化得到热稳定超氧化物歧化酶(SOD),纯化倍数13.4,回收率11.1%,酶比活力达3354 U/mg.SOD对KCN和H₂O₂不敏感性,说明来自Geobacillus sp. EPT3的超氧化物歧化酶为Mn-SOD类型.SOD为同聚二聚体,亚基分子量为26.0 kDa.该酶在50℃和60℃下有高的热稳定性.在检测条件下,SOD在一些抑制剂和变性剂存在时相对稳定,例如β-ME、DTT、PMSF、尿素和盐酸胍.Geobacillus sp. EPT3超氧化物歧化酶在pH 5.0~11.0的广范围内有好的稳定性,能耐受极酸和极碱的变性条件,有利于它的工业应用.
- Geobacillussp. /
- 超氧化物歧化酶 /
- 热稳定性 /
- 纯化
Abstract: Thermostable SOD is a promising enzyme in biotechnological applications. In the present study, thermophile Geobacillus sp. EPT3 was isolated from a deep-sea hydrothermal field in the East Pacific. A thermostable superoxide dismutase (SOD) from this strain was purified to homogeneity by steps of fractional ammonium sulfate precipitation, DEAE-Sepharose chromatography, and Phenyl-Sepharose chromatography. SOD was purified 13.4 fold to homogeneity with a specific activity of 3 354 U/mg and 11.1% recovery. SOD from Geobacillus sp. EPT3 was of the Mn-SOD type, judged by the insensitivity of the enzyme to both KCN and H₂O₂. SOD was determined to be a homodimer with monomeric molecular mass of 26.0 kDa. It had high thermostability at 50℃ and 60℃. At tested conditions, SOD was relatively stable in the presence of some inhibitors and denaturants, such as β-mercaptoethanol (β-ME), dithiothreitol (DTT), phenylmethylsulfonyl fluoride (PMSF), urea, and guanidine hydrochloride. Geobacillus sp. EPT3 SOD showed striking stability across a wide pH range from 5.0 to 11.0. It could withstand denaturants of extremely acidic and alkaline conditions, which makes it useful in the industrial applications.
- Geobacillus sp. /
- superoxide dismutase /
- thermostability /
- purification

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深海嗜热菌Geobacillus sp. EPT3锰超氧化物歧化酶的纯化与生物化学鉴定

doi: 10.1007/s13131-014-0534-2

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Purification and biochemical characterization of manganesecontaining superoxide dismutase from deep-sea thermophile Geobacillus sp. EPT3

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