CN102277317B - Lipoxygenase-producing pseudomonas aeruginosa and screening method and application thereof - Google Patents
Lipoxygenase-producing pseudomonas aeruginosa and screening method and application thereof Download PDFInfo
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- CN102277317B CN102277317B CN 201110212308 CN201110212308A CN102277317B CN 102277317 B CN102277317 B CN 102277317B CN 201110212308 CN201110212308 CN 201110212308 CN 201110212308 A CN201110212308 A CN 201110212308A CN 102277317 B CN102277317 B CN 102277317B
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Abstract
The invention discloses a lipoxygenase-producing pseudomonas aeruginosa and a screening method and an application thereof, and belongs to the biotechnology field. According to the invention, a soil sample is coated on a screening plate which contains linoleic acid and beta-carotene for primary screening, and bacterial strains with a color-changing circle are selected for secondary screening in a shake flask. A strain of lipoxygenase-producing pseudomonas aeruginosa is successfully screened. The screening method of the invention can effectively increase the efficiency of screening of lipoxygenase-producing strains, and is applicable to seed selection of lipoxygenase-producing strains.
Description
Technical field
The present invention relates to a Pseudomonas aeruginosa strain and screening method thereof and application, belong to biological technical field.
Background technology
Pseudomonas aeruginosa (
P.Aeruginosa) former title Pseudomonas aeruginosa.Extensive in distributed in nature, be one of modal bacterium of existing in the soil.Obligate aerobe, 25 ~ 42 ℃ of growth temperature ranges, optimum growth temperature are 25 ~ 30 ℃.In recent years, have the research report to point out, Pseudomonas aeruginosa contains fat oxygenase gene.
Lipoxygenase (Lipoxygenase, EC1.13.11.12 are called for short LOX) belongs to oxydo-reductase, can single-minded catalysis contain cis, cis-1, polybasic unsaturated fatty acid and the ester thereof of 4-pentadiene structure, make its pass through minute within oxygenation, form the hydroperoxide with conjugated double bond.
Lipoxygenase claims again the unsaturated fatty acids oxydo-reductase, be present in clover, pea, French beans, etc. in the leguminous plants, also have in radish and the potato, but the strongest with the activity in the soybean.By the lipoxygenase of crystallization mostly, relative molecular weight is 102000, and iso-electric point is 5.14.Optimal pH can not simply determine, but is subjected to about the solvability of substrate, general pH 6.10-3.10, and optimum temperuture is between 20-30 ℃.But lipoxygenase catalytic molecular oxygen to having the grease generation oxidation of the two keys of pentadiene Isosorbide-5-Nitrae in the flour, forms hydroperoxide.(SH) form disulfide linkage (S-S-) and can the induced protein molecule aggregation, makes protein molecule become larger, thereby strengthens the effect of gluten the sulfydryl of hydroperoxide oxidized protein molecule.Lipoxygenase can be destroyed by coupled reaction the double bond structure of Radix Dauci Sativae, thereby plays bleaching flour, improves the effect of flour color and luster.And the superoxide that lipoxidase enzyme catalysis linolic acid generates can improve the fragrance of bread, is the bread flavouring.This shows the effect that lipoxygenase has strong muscle concurrently and brightens, the consumption that can be used for alternative gluten-strengthening agent potassium bromate and reduce the SYNTHETIC OPTICAL WHITNER benzoyl peroxide.
Summary of the invention
Technical problem to be solved by this invention provides a kind of Pseudomonas aeruginosa that produces lipoxygenase, this bacterial strain was on May 30th, 2011, be preserved in Chinese Typical Representative culture collection center, deposit number is CCTCC NO:M 2011185, address: China, Wuhan, Wuhan University, its taxonomy called after Pseudomonas aeruginosa BBE (
Pseudomonas aeruginosaBBE).
Another technical problem to be solved by this invention provides a kind of method of screening the Pseudomonas aeruginosa that produces lipoxygenase.
The technical scheme that addresses the above problem is: after pedotheque dilutes with sterilized water near the grease factory, coat on the LB flat board that contains 0.5% linolic acid and 0.5% β-carotene.Select the bacterial strain that can produce the variable color circle, be inoculated in the liquid nutrient medium that contains 1% glucose and 2% oleic acid, cultivate for 30 ℃ and measure respectively the activity of fatty oxygenase that born of the same parents are outer and born of the same parents are interior after 24h hour.
The present invention has following advantage:
1) screens the Pseudomonas aeruginosa that obtains and to produce lipoxygenase.
2) the simple and successful of selection provided by the invention.
Embodiment
The screening method of embodiment 1 Pseudomonas aeruginosa (CCTCC NO:M 2010292)
Gather soil from grease factory, with 1g soil and 10ml sterilized water mixing, use filter paper filtering, filtrate coating contained the soil diluent is coated on the LB flat board that contains 0.005% linolic acid and 0.005% β-carotene, behind 30 ℃ of cultivation 20h, there is the bacterium colony of variable color circle on the LB flat board that contains 0.005% linolic acid and 0.005% β-carotene, to rule around the choosing colony.Selecting line has single bacterium colony of variable color circle, is inoculated in the GL substratum, cultivates 24h for 30 degrees centigrade, and the centrifugal 2min of 1000rpm collects supernatant liquor and surveys activity of fatty oxygenase; Thalline 50mM, the Tris-HCl damping fluid of pH7.0 redissolve and use ultrasonication, and the centrifugal 2min of shell-broken liquid 1000rpm gets supernatant liquor and surveys activity of fatty oxygenase.
The lipoxidase enzyme activity is 300U/mL.
Although the present invention with preferred embodiment openly as above; but it is not to limit the present invention, any person skilled in the art, without departing from the spirit and scope of the present invention; all can do various changes and modification, so protection scope of the present invention should be with being as the criterion that claims were defined.
Claims (2)
1. one kind is produced lipoxygenase Pseudomonas aeruginosa BBE (Pseudomonas aeruginosa) BBE, is preserved in Chinese Typical Representative culture collection center, and deposit number is CCTCC NO:M 2011185.
2. the application of the described Pseudomonas aeruginosa of claim 1 in the lipoxygenase fermentative production.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 201110212308 CN102277317B (en) | 2011-07-28 | 2011-07-28 | Lipoxygenase-producing pseudomonas aeruginosa and screening method and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 201110212308 CN102277317B (en) | 2011-07-28 | 2011-07-28 | Lipoxygenase-producing pseudomonas aeruginosa and screening method and application thereof |
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| Publication Number | Publication Date |
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| CN102277317A CN102277317A (en) | 2011-12-14 |
| CN102277317B true CN102277317B (en) | 2013-01-02 |
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Families Citing this family (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102994465B (en) * | 2012-12-27 | 2015-03-25 | 江南大学 | Lipoxygenase with increased enzymatic activity and heat stability and construction method of lipoxygenase |
| CN104630164A (en) * | 2015-02-16 | 2015-05-20 | 大连大学 | Method for producing low-temperature lipoxygenase by fermenting marine microorganisms |
| CN104630165A (en) * | 2015-02-16 | 2015-05-20 | 大连大学 | Method for producing low-temperature lipoxygenase employing microbial fermentation |
| WO2022191638A1 (en) * | 2021-03-10 | 2022-09-15 | 이화여자대학교 산학협력단 | Lipoxygenase-based recombinant microorganisms, and method for preparing hydroxy fatty acids and secondary fatty alcohols using same |
| CN116042446B (en) * | 2022-09-30 | 2024-04-30 | 哈尔滨工业大学 | Rhamnolipid producing strain for oil-stone separation of waste asphalt mixture and application thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2002086114A1 (en) * | 2001-04-20 | 2002-10-31 | Novozymes A/S | Lipoxygenase |
| CN101643740A (en) * | 2009-08-28 | 2010-02-10 | 南京农业大学 | Anabaena PCC7120 fat oxygenase gene |
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2011
- 2011-07-28 CN CN 201110212308 patent/CN102277317B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2002086114A1 (en) * | 2001-04-20 | 2002-10-31 | Novozymes A/S | Lipoxygenase |
| CN101643740A (en) * | 2009-08-28 | 2010-02-10 | 南京农业大学 | Anabaena PCC7120 fat oxygenase gene |
Non-Patent Citations (1)
| Title |
|---|
| Jae-Han Bae等.Thermostable lipoxygenase is a key enzyme in the conversion of linoleic acid to trihydroxy-octadecenoic acid by Pseudomonas aeruginosa PR3.《Biotechnology and Bioprocess Engineering》.2010,第15卷(第6期),1022-1030. * |
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| CN102277317A (en) | 2011-12-14 |
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Address after: No. 258 Wuxing Jiayuan, Liangxi District, Wuxi City, Jiangsu Province Patentee after: Jiangnan University Address before: 1800 No. 214122 Jiangsu city of Wuxi Province Li Lake Avenue Patentee before: Jiangnan University |