CN104328078B - Brevibacillus borstelensis for producing neutral protease - Google Patents
Brevibacillus borstelensis for producing neutral protease Download PDFInfo
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- CN104328078B CN104328078B CN201410668536.0A CN201410668536A CN104328078B CN 104328078 B CN104328078 B CN 104328078B CN 201410668536 A CN201410668536 A CN 201410668536A CN 104328078 B CN104328078 B CN 104328078B
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Abstract
The invention relates to brevibacillus borstelensis for producing neutral protease. The invention provides brevibacillus borstelensis for producing neutral protease. The brevibacillus borstelensis for producing neutral protease disclosed by the invention is brevibacillus borstelensis HWyb1312 which is preserved in China General Microbiological Culture Collection Center with the preservation number of CGMCC No. 8717. The brevibacillus borstelensis HWyb1312 disclosed by the invention has a high-temperature resistance characteristic, and can survive at 70 DEG C and metabolize neutral protease.
Description
Technical field
The present invention relates to Potsdam bacillus brevis of one plant of product protease.
Background technology
Neutral protease is usually obtained by the fermented extraction of bacillus subtilises, belongs to a kind of restriction endonuclease, can be used for
Various proteolysis are processed.In uniform temperature (optimum temperature：45 DEG C～50 DEG C), pH value (stablize pH：5.5～8.5) under, in
Macro-molecular protein can be hydrolyzed to the products such as aminoacid by property protease, can be used for the hydrolysis of animal and vegetable protein.Due to neutral egg
The macro-molecular protein of animal and plant can be hydrolyzed into small-molecular peptides or aminoacid by white enzyme, be beneficial to effective absorption of protein
And utilization, its hydrolyzed solution AN% height, degree of hydrolysis is high, and local flavor is good, mainly for the production of senior flavoring agent and food enrichment,
And various animal origin extract produces feature bone, meat extract (ossein), Aquatic product extract, peptone, peptide etc..Remove
Outside this, neutral protease is baking industry, brewing industry, medical industry, textile industry, leather industry, soybean separation protein white collar
Domain, yeast extract, the production field of yeast extract are also all widely used；And, also used as feed ingredient.
Content of the invention
The present invention provides Potsdam bacillus brevis of one plant of product neutral protease, and the acquisition that increased neutral protease comes
Source.
Potsdam bacillus brevis that the present invention produces neutral protease are Potsdam bacillus brevis (Brevibacillus
Borstelensis) HWyb1312, is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, preserving number
For CGMCC No.8717.
Potsdam of the present invention bacillus brevis (Brevibacillus borstelensis) HWyb1312 is gram sun
Property bacterium, the morphological characteristic of thalline is rod-short, and cell size is (0.3 μm～0.4 μm) × (2 μm～7 μm), has significantly recessed
Fall into, peritrichouses, central spore, motion, as shown in Figure 1；Bacterium colony is rounded, and color is milky, and smooth surface is opaque,
Not raised, as shown in Figure 2.
The most suitable growth of Potsdam of the present invention bacillus brevis (Brevibacillus borstelensis) HWyb1312
Temperature is 50 DEG C～60 DEG C, and the pH value being best suitable for growing environment is 6.5～7.0.
Potsdam of the present invention bacillus brevis (Brevibacillus borstelensis) HWyb1312 oxydase reaction
It is negative, gelatin experimental result is positive, Starch Hydrolysis experimental result is negative, nitrate reduction reaction is positive, and V-P reacts
It is negative, glucose response is positive, citrate reaction is positive, arabinose reaction is negative, xylose reaction is negative.
GN plate using the identification of Mei Liai VITEK 2Compact automatic bacterial and Analysis of Drug Susceptibility system is short to Potsdam
The biochemical reaction of bacillus cereuss (Brevibacillus borstelensis) HWyb1312 is identified.The short spore in Potsdam
The biochemical reaction result of bacillus (Brevibacillus borstelensis) HWyb1312 is as shown in table 1.
Table 1
| BXYL | - | LysA | - | AspA | + | LeuA | + |
| PheA | + | BGAL | - | PyrA | - | AGAL | - |
| AlaA | + | TyrA | + | APPA | - | CDEX | - |
| dGAL | - | GLYG | - | INO | - | ELLM | - |
| MdX | - | AMAN | - | MTE | - | GIyA | + |
| dMNE | - | dMLZ | - | NAG | - | PLE | - |
| IRHA | - | BMAN | - | PHC | + | PVATE | - |
| AGLU | - | dTAG | - | dTRE | - | INU | - |
| dGLU | - | dRIB | - | PSCNa | - | NaCl 6.5% | - |
| KAN | + | OLD | - | ESC | + | TTZ | - |
| POLYB_R | - | ProA | - | BNAG | - | MdG | - |
| dMAN | - | BGLU | - |
Potsdam of the present invention bacillus brevis (Brevibacillus borstelensis) HWyb1312 passes through 16S
RDNA gene order compares analysis, the 16S rDNA with Potsdam bacillus brevis (Brevibacillus borstelensis)
The homology highest of gene order, similarity is 100%, by with reference to morphological features, growth conditionss, Physiology and biochemistry mirror
Fixed, it is defined as Potsdam bacillus brevis (Brevibacillus borstelensis).
The present invention produces bacillus brevis Potsdam, Potsdam bacillus brevis of neutral protease
(Brevibacillus borstelensis) HWyb1312 has high-temperature stability, remains to survive under the conditions of 70 DEG C, and energy
Metabolism goes out neutral protease, and under the conditions of 55 DEG C, in its fermentation liquid, the enzyme activity of neutral protease is up to 57U/mL.The short bud in Potsdam
Spore bacillus (Brevibacillus borstelensis) HWyb1312 can be used for≤and 70 DEG C of hot fermentation produces neutral protein
Enzyme.
Due to Potsdam of the present invention bacillus brevis (Brevibacillus borstelensis) HWyb1312 have resistance to
High temperature and the double grading producing neutral protease, can be used for the fields such as During High-Temperature Composting, the high temperature sewage disposal of fowl and animal excrement, thus
Efficient degradation protein therein.
Potsdam of the present invention bacillus brevis (Brevibacillus borstelensis) HWyb1312 is that Potsdam is short
Bacillus cereuss (Brevibacillus borstelensis), belong to bacillus (Bacillus)；It is preserved in Chinese micro- life
Thing culture presevation administration committee common micro-organisms center (CGMCC), preservation address is Yard 1, BeiChen xi Road, Chaoyang District, Beijing City
No. 3, preservation date is on January 10th, 2014, and preserving number is CGMCC No.8717.
Brief description
Fig. 1 is that Potsdam bacillus brevis (Brevibacillus borstelensis) HWyb1312 thalline microscope is seen
Examine figure.
Fig. 2 is that Potsdam bacillus brevis (Brevibacillus borstelensis) HWyb1312 bacterium colony observes figure.
Specific embodiment
Technical solution of the present invention is not limited to act specific embodiment set forth below, also includes between each specific embodiment
Combination in any.
Specific embodiment one：Potsdam bacillus brevis that present embodiment produces neutral protease are the short spore in Potsdam
Bacillus (Brevibacillus borstelensis) HWyb1312, is preserved in China Committee for Culture Collection of Microorganisms general
Logical microorganism center, preserving number is CGMCC No.8717.
Present embodiment Potsdam bacillus brevis (Brevibacillus borstelensis) HWyb1312 is isolatable from
Qiqihaer City Gannan County compost fertilizer.
Screening is carried out according to the following steps：
Weigh 10.0g compost fertilizer to put in the triangular flask of the 100ml sterilized water containing bead, in 200r/min concussion
30min, makes bacteria suspension；Take 1ml suspension to be added in 100mL enrichment medium, shake under conditions of 55 DEG C, 180r/min
Culture 18h～20h, carries out gradient dilution afterwards, coats in casein screening culture medium, is put in culture 48h in 55 DEG C of environment, chooses
Select the bacterial strain producing transparent circle, obtain present embodiment Potsdam bacillus brevis (Brevibacillus
borstelensis)HWyb1312.In enrichment medium 1000ml contain casein 2.0g, yeast extract 1.0g, glucose 10g and
The water of surplus, pH value is 7.0.Casein screening culture medium by peptone 10.0g, glucose 1.0g, NaCl 5.0g,
CaCl20.1g, L-Tyr 0.1g, casein 5.0g, agar 18.0g and H2O 1000ml makes, and pH value is 7.0.
Available present embodiment Potsdam bacillus brevis (Brevibacillus borstelensis) HWyb1312
Produce neutral protease, its fermentation medium is by Carnis Bovis seu Bubali cream 3.0g, peptone 10.0g, NaCl 5.0g and H2O 1000ml system
Become, pH value is 7.0.
Present embodiment Potsdam bacillus brevis (Brevibacillus borstelensis) HWyb1312 can be used for
The During High-Temperature Composting of fowl and animal excrement, decomposable asymmetric choice net macro-molecular protein therein.
Specific embodiment two：Present embodiment Potsdam bacillus brevis (Brevibacillus borstelensis)
HWyb1312 is inoculated in beef extract-peptone fermentation medium with 1% inoculum concentration, in the condition of different temperatures of 220r/min
Lower culture 72h, frozen centrifugation 30min under the conditions of 10000r/min, 4 DEG C afterwards, collect supernatant and adopt Folin- phenol to develop the color
Method measures the neutral protease activity of crude enzyme liquid.Under the conditions of pH value is 7.2, hydrolyzed casein per minute produces 1 μ g tyrosine and is 1
Enzyme-activity unit (U/mL).
In condition of different temperatures bottom fermentation culture fluid, the enzyme activity of neutral protease is as shown in table 1.
Table 1
| Fermentation temperature (DEG C) | Neutral protease vigor (U/mL) |
| 50 | 51.6 |
| 55 | 57 |
| 60 | 52.9 |
| 65 | 46.3 |
| 70 | 38.2 |
Experiment proves that Potsdam bacillus brevis (Brevibacillus borstelensis) HWyb1312 sends out at 70 DEG C
Still active under the conditions of ferment, and neutral protease can be produced.
Claims (1)
1. Potsdam bacillus brevis of one plant of product neutral protease, it is Potsdam bacillus brevis (Brevibacillus
Borstelensis) HWyb1312, is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, preserving number
For CGMCC No.8717.
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Families Citing this family (7)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105211497A (en) * | 2015-10-22 | 2016-01-06 | 中国科学院微生物研究所 | Novel composite probiotic feed additive and application thereof |
| CN110760466B (en) * | 2019-11-18 | 2021-04-20 | 山东隆科特酶制剂有限公司 | Bacillus subtilis for producing high-temperature-resistant neutral protease and application thereof |
| CN112195115B (en) * | 2020-08-13 | 2023-03-17 | 长江大学 | Brevibacillus borstelensis, preparation, method for producing surfactant and application |
| CN114164145B (en) * | 2021-11-23 | 2022-09-02 | 福建农林大学 | Brevibacillus borstelensis, neutral protease and application thereof |
| CN114456982B (en) * | 2022-03-03 | 2023-09-26 | 青岛蔚蓝赛德生物科技有限公司 | Brevibacillus brevis and application thereof in degrading or digesting sludge |
| CN116179421A (en) * | 2022-12-15 | 2023-05-30 | 黄鹤楼酒业有限公司 | Brevibacillus borstele HIL-001 and application thereof |
| CN117265020B (en) * | 2023-11-21 | 2024-02-20 | 烟台融科生物科技有限公司 | Method for co-producing pig bone essence and bone collagen active peptide by using pig bone |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101407777A (en) * | 2008-12-01 | 2009-04-15 | 大庆油田有限责任公司 | Potsdam Bacillus brevis and use thereof |
| CN103013877A (en) * | 2012-12-14 | 2013-04-03 | 中国科学院广州地球化学研究所 | Brevibacillus borstelensis strain having capability of degrading thioanisole and application thereof |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101407777A (en) * | 2008-12-01 | 2009-04-15 | 大庆油田有限责任公司 | Potsdam Bacillus brevis and use thereof |
| CN103013877A (en) * | 2012-12-14 | 2013-04-03 | 中国科学院广州地球化学研究所 | Brevibacillus borstelensis strain having capability of degrading thioanisole and application thereof |
Non-Patent Citations (1)
| Title |
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| Optimization of Mineral sources on α-amylase Production by Brevibacillus borstelensis R1in Submerged Fermentationx-xbitm;K.SuribabuBly et al;《International Journal of Engineering Sciences & Research Technology》;20140731;第3卷(第7期);926-934 * |
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Inventor after: Yin Bo Inventor after: Wu Haoqiong Inventor after: Niu Yanbo Inventor after: Yang Xusheng Inventor after: Liu Leiming Inventor after: Wang Yuwen Inventor after: Cao Yabin Inventor after: Sun Yufeng Inventor after: Guo Lishu Inventor after: An Qi Inventor after: Fan Chuan Inventor after: Wang Huimin Inventor after: Zhang Xin Inventor after: Chi Jundao Inventor before: Yin Bo Inventor before: Wu Haoqiong Inventor before: Niu Yanbo Inventor before: Yang Xusheng Inventor before: Liu Leiming Inventor before: Wang Yuwen Inventor before: Cao Yabin Inventor before: Sun Yufeng Inventor before: Guo Lishu Inventor before: An Qi Inventor before: Fan Chuan Inventor before: Wang Huimin Inventor before: Zhang Xin Inventor before: Chi Jundao |
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