CN102071156B - Bacillus licheniformis capable of producing beta-mannanase - Google Patents
Bacillus licheniformis capable of producing beta-mannanase Download PDFInfo
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- CN102071156B CN102071156B CN2009100734733A CN200910073473A CN102071156B CN 102071156 B CN102071156 B CN 102071156B CN 2009100734733 A CN2009100734733 A CN 2009100734733A CN 200910073473 A CN200910073473 A CN 200910073473A CN 102071156 B CN102071156 B CN 102071156B
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- bacillus licheniformis
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- mannanase
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Abstract
The invention discloses bacillus licheniformis capable of producing beta-mannanase, which relates to bacillus licheniformis. The invention solves the low enzymatic activity problem of the conventional production of beta-mannanase by fungi. The bacillus licheniformis capable of producing beta-mannanase belongs to a new bacillus licheniformis variety, and was collected in China Center for Type Culture Collection at Wuhan University in Wuhan on November 29th, 2009, with a collection number of CGTTC No.M209288; the bacillus licheniformis capable of producing beta-mannanase is Gram-positive and anaerobic bacillus, is 0.5 to 0.8-micrometer wide, 1.5 to 2.0-micrometer long and does not have flagella and mesic elliptic spores. The beta-mannanase produced by the bacillus licheniformis is acidic mannanase, as is proved by a test. The molecular weight of the beta-mannanase is about 38kD. The beta-mannanase has higher high temperature resistance activity, and the highest activity of the beta-mannanase is 5168U/mL.
Description
Technical field
The present invention relates to a kind of Bacillus licheniformis.
Background technology
'beta '-mannase is hydrolysis 1, and the inscribe lytic enzyme of 4-β-D-mannopyranose main chain belongs to the semicellulose enzyme, and the effect substrate mainly is polygalactomannan, glucomannan, gala glucomannan and mannosans.Being used widely in many-sides such as food, medicine, papermaking, textile printing and dyeing, oil production and biological study technology, is a kind of very important instrument enzyme.Mostly the employed bacterial strain of present industrial production is fungi, and enzyme work is merely 2300U/mL.
Summary of the invention
The objective of the invention is to solve and utilize fungi generation 'beta '-mannase to have enzyme low problem alive now, and a kind of Bacillus licheniformis that produces 'beta '-mannase is provided.
Bacillus licheniformis (Bacilluslicheniformis) HDGLJT-01 that the present invention can produce 'beta '-mannase belongs to Bacillus licheniformis (Bacillus) novel species through evaluation; Be preserved in Chinese typical culture collection center; The preservation address is a Wuhan City Wuhan University; Preservation date is on November 29th, 2009, and deposit number is CGTCC No.M209288; The Bacillus licheniformis that can produce 'beta '-mannase is the Gram-positive facultative Bacteroides nodosus, and wide is 0.5～0.8 μ m, and length is 1.5～2.0 μ m; Atrichia, the oval gemma of middle life; Salt tolerant 10%, growth temperature range are 30 ℃～55 ℃, the single arrangement of thalline or be chain; The Bacillus licheniformis that can the produce 'beta '-mannase bacterium colony of on solid medium, growing is an oyster white, circle, and the edge is irregular, and surface irregularity is opaque, and colony diameter is 0.5～1.5mm.
The present invention can produce that the Bacillus licheniformis catalase test of 'beta '-mannase is positive, the starch hydrolysis experiment is positive, the gelatin hydrolysis test is positive, the Fu Pu test is positive, D-glucose produces that the acid test is positive, nitrate reduction test is positive, L-arabinose produce acid, D-wood sugar produce acid, D-N.F,USP MANNITOL produce acid, Citrate trianion utilization, propionic salt utilization, glucose aerogenesis test negative, tyrosine hydrolysis test is negative, indole test is negative.
The 16S rDNA sequence length that the present invention can produce the Bacillus licheniformis of 'beta '-mannase is 1605bp; The BLAST compare of analysis can know that the kind the most close with it is Bacillus licheniformis Bacillus licheniformis (DQ020262), and similarity is 99.8%.
The present invention can produce the Bacillus licheniformis of 'beta '-mannase, and the 'beta '-mannase that is produced is acid mannase, and molecular weight is about 38kD; Suitable temperature of reaction is 50 ℃～60 ℃; Optimum temperuture is 55 ℃, and behind the insulation 30min, the temperature T 1/2 of enzyme half inactivation is 70 ℃; Suitable reaction pH value is 4.5～6.0, poor stability when good stability when the pH value is acid, neutral and meta-alkalescence, and this enzyme shows high temperature resistant preferably activity, and enzymatic activity high reaches 5168U/mL.
Bacillus licheniformis belongs to Bacillus licheniformis (Bacillus), is preserved in Chinese typical culture collection center, and the preservation address is a Wuhan City Wuhan University, and preservation date is on November 29th, 2009, and deposit number is CGTCC No.M209288.
Embodiment
Embodiment one: Bacillus licheniformis (Bacillus licheniformis) HDGLJT-01 that this embodiment can produce 'beta '-mannase belongs to Bacillus licheniformis (Bacillus) novel species through evaluation; Be preserved in Chinese typical culture collection center; The preservation address is a Wuhan City Wuhan University; Preservation date is on November 29th, 2009, and deposit number is CGTCC No.M209288; The Bacillus licheniformis that can produce 'beta '-mannase is the Gram-positive facultative Bacteroides nodosus, and wide is 0.5～0.8 μ m, and length is 1.5～2.0 μ m; Atrichia, the oval gemma of middle life; Salt tolerant 10%, growth temperature range are 30 ℃～55 ℃, the single arrangement of thalline or be chain; The Bacillus licheniformis that can the produce 'beta '-mannase bacterium colony of on solid medium, growing is an oyster white, circle, and the edge is irregular, and surface irregularity is opaque, and colony diameter is 0.5～1.5mm.
The 16S rDNA sequence length that this embodiment can produce the Bacillus licheniformis of 'beta '-mannase is 1605bp; The BLAST compare of analysis can know that the kind the most close with it is Bacillus licheniformis Bacilluslicheniformis (DQ020262), and similarity is 99.8%.
With reference to " uncle's outstanding Bacteria Identification handbook the 8th edition is carried out Physiology and biochemistry with " common bacteria system identification handbook " to the Bacillus licheniformis that can produce 'beta '-mannase and identified: the Bacillus licheniformis catalase test that can produce 'beta '-mannase is positive, the starch hydrolysis experiment is positive, the gelatin hydrolysis test is positive, the Fu Pu test is positive, D-glucose produces that the acid test is positive, nitrate reduction test is positive, L-arabinose produce acid, D-wood sugar produce acid, D-N.F,USP MANNITOL produce acid, Citrate trianion utilization, propionic salt utilization, glucose aerogenesis test negative, tyrosine hydrolysis test is negative, indole test is negative.
This embodiment can produce the Bacillus licheniformis of 'beta '-mannase, and the 'beta '-mannase that is produced is acid mannase, and molecular weight is about 38kD; Suitable temperature of reaction is 50 ℃～60 ℃; Optimum temperuture is 55 ℃, and behind the insulation 30min, the temperature T 1/2 of enzyme half inactivation is 70 ℃; Suitable reaction pH value is 4.5～6.0, poor stability when good stability when the pH value is acid, neutral and meta-alkalescence, and this enzyme shows high temperature resistant preferably activity, and enzymatic activity high reaches 5168U/mL.
Embodiment two: screening obtains Bacillus licheniformis (Bacillus licheniformis) HDGLJT-01 that this embodiment can produce 'beta '-mannase from main biological stage retting liquid.Screening is carried out according to the following steps: get main biological stage retting liquid 2mL and place the 100mL enrichment medium to cultivate, enrichment culture is 2 days in 37 ℃, the environment of 160r/min; Be inoculated in then on the Rhizoma amorphophalli powder isolation medium, under 37 ℃ of conditions, cultivated 2 days, obtain single bacterium colony, be Bacillus licheniformis.
Enrichment medium is by 5g Rhizoma amorphophalli powder, 1g K in this embodiment
2HPO
43H
2O and 1000mL zero(ppm) water are formed, and regulate pH value 7.2, at 121 ℃ of 15min that sterilize down.
The Rhizoma amorphophalli powder isolation medium is by 5g Rhizoma amorphophalli powder, 10g peptone, 2g yeast extract paste, 0.2g MgCl in this embodiment
26H
2O, 1g K
2HPO
43H
2O, 20g agar powder and 1000mL zero(ppm) water are formed, and regulate pH value 7.2, at 121 ℃ of 15min that sterilize down.
Gained Bacillus licheniformis in this embodiment, the 'beta '-mannase that is produced is acid mannase through test, molecular weight is about 38kD; Suitable temperature of reaction is 50 ℃～60 ℃; Optimum temperuture is 55 ℃, and behind the insulation 30min, the temperature T 1/2 of enzyme half inactivation is 70 ℃; Suitable reaction pH value is 4.5～6.0, poor stability when good stability when the pH value is acid, neutral and meta-alkalescence, and this enzyme shows high temperature resistant preferably activity, and enzymatic activity high reaches 5168U/mL.
Sequence table
< 110>Heilongjiang University
< 120>a kind of Bacillus licheniformis that produces 'beta '-mannase
<160>1
<210>1
<211>1605
<212>DNA
< 213>can produce Bacillus licheniformis (Bacillus licheniformis) HDGLJT-01 of 'beta '-mannase
<220>
< 223>can produce Bacillus licheniformis (Bacillus licheniformis) the HDGLJT-01 16SrDNA gene order of 'beta '-mannase
<400>1
gctcccggcc?gccatggcgg?ccgcgggaat?tcgattagag?tttgatcctg?gctcaggacg 60
aacgctggcg?gcgtgcctaa?tacatgcaag?tcgagcggac?agatgggagc?ttgctccctg?120
atgttagcgg?tggacgggtg?agtaacacgt?gggtaacctg?cctgtaagac?tgggataact?180
ccgggaaacc?ggggctaata?ccggatggtt?gtttgaaccg?catggttcaa?acataaaagg?240
tggcttcggc?taccacttac?agatggaccc?gcggcgcatt?agctagttgg?tgaggtaacg?300
gctcaccaag?gcaacgatgc?gtagccgacc?tgagagggtg?atcggccaca?ctgggactga?360
gacacggccc?agactcctac?gggaggcagc?agtagggaat?cttccgcaat?ggacgaaagt?420
ctgacggagc?aacgccgcgt?gagtgatgaa?ggttttcgga?tcgtaaagct?ctgttgttag?480
ggaagaacaa?gtaccgttcg?aatagggcgg?taccttgacg?gtacctaacc?agaaagccac?540
ggctaactac?gtgccagcag?ccgcggtaat?acgtaggtgg?caagcgttgt?ccggaattat?600
tgggcgtaaa?gggctcgcag?gcggtttctt?aagtctgatg?tgaaagcccc?cggctcaacc?660
ggggagggtc?attggaaact?ggggaacttg?agtgcagaag?aggagagtgg?aattccacgt?720
gtagcggtga?aatgcgtaga?gatgtggagg?aacaccagtg?gcgaaggcga?ctctctggtc?780
tgtaactgac?gctgaggagc?gaaagcgtgg?ggagcgaaca?ggattagata?ccctggtagt?841
ccacgccgta?aacgatgagt?gctaagtgtt?agggggtttc?cgccccttag?tgctgcagct?900
aacgcattaa?gcactccgcc?tggggagtac?ggtcgcaaga?ctgaaactca?aaggaattga?6902
cgggggcccg?cacaagcggt?ggagcatgtg?gtttaattcg?aagcaacgcg?aagaacctta?1020
ccaggtcttg?acatcctctg?acaatcctag?agataggacg?tccccttcgg?gggcagagtg?1080
acaggtggtg?catggttgtc?gtcagctcgt?gtcgtgagat?gttgggttaa?gtcccgcaac?1140
gagcgcaacc?cttgatctta?gttgccagca?ttcagttggg?cactctaagg?tgactgccgg?1200
tgacaaaccg?gaggaaggtg?gggatgacgt?caaatcatca?tgccccttat?gacctgggct?1260
acacacgtgc?tacaatggac?agaacaaagg?gcagcgaaac?cgcgaggtta?agccaatccc?1320
acaaatctgt?tctcagttcg?gatcgcagtc?tgcaactcga?ctgcgtgaag?ctggaatcgc?1380
tagtaatcgc?ggatcagcat?gccgcggtga?atacgttccc?gggccttgta?cacaccgccc?1440
gtcacaccac?gagagtttgt?aacacccgaa?gtcggtgagg?taacctttta?ggagccagcc?1500
gccgaaggtg?ggacagatga?ttggggtgaa?gtcgtaacaa?ggtagccgta?atcactagtg?1560
aattcgcggc?cgcctgcagg?tcgaccatat?gggagagctc?ccaac 1605
Claims (1)
1. Bacillus licheniformis that can produce 'beta '-mannase; Bacillus licheniformis (Bacillus licheniformis) HDGLJT-01 that it is characterized in that producing 'beta '-mannase belongs to Bacillus licheniformis (Bacillus licheniformis) novel species through evaluation; Be preserved in Chinese typical culture collection center; The preservation address is a Wuhan City Wuhan University, and preservation date is on November 29th, 2009, and deposit number is CGTCC No.M209288; Its characteristics are the Gram-positive facultative Bacteroides nodosus, and wide is 0.5～0.8 μ m, and length is 1.5～2.0 μ m, atrichia, the oval gemma of middle life, and growth temperature range is 30 ℃～55 ℃, the single arrangement of thalline or be chain; The growth bacterium colony is an oyster white on solid medium, circle, and the edge is irregular, and surface irregularity is opaque, and colony diameter is 0.5～1.5mm.
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Families Citing this family (11)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN103184206B (en) * | 2011-08-16 | 2015-04-15 | 青岛蔚蓝生物集团有限公司 | Mannase and mutants thereof |
| CN103740682B (en) * | 2014-01-17 | 2016-01-06 | 北京科为博生物科技有限公司 | A kind of high temperature resistant acidic 'beta '-mannase Man-L30 and gene thereof and application |
| CN104962538A (en) * | 2015-07-13 | 2015-10-07 | 黑龙江大学 | Method for producing beta-mannase by bacillus germs |
| CN104962542B (en) * | 2015-07-13 | 2018-09-14 | 黑龙江大学 | A kind of method of sporosarcina production pectase and mannase |
| CN104946613B (en) * | 2015-07-13 | 2018-09-14 | 黑龙江大学 | A kind of method of Sporosarcina bacterium production 'beta '-mannase |
| CN105621630B (en) * | 2016-01-18 | 2019-02-01 | 黑龙江大学 | A kind of decoloration 'beta '-mannase and application thereof |
| CN105936899B (en) * | 2016-01-19 | 2019-05-17 | 黑龙江大学 | A kind of bacillus licheniformis produces the feed process of 'beta '-mannase |
| CN109825451A (en) * | 2019-01-25 | 2019-05-31 | 牡丹江医学院 | A kind of 'beta '-mannase superior strain and its application in dye decolored |
| CN111040966B (en) * | 2019-12-23 | 2022-06-24 | 河北科技大学 | Bacillus licheniformis KD-1, beta-mannase produced by same and application thereof |
| CN113186127A (en) * | 2021-04-20 | 2021-07-30 | 中国石油大学(华东) | Guanidine gum fracturing fluid degrading bacterium, culture method and application thereof |
| CN115232773B (en) * | 2022-08-05 | 2023-12-19 | 武汉市四圣元生物科技有限责任公司 | Bacillus licheniformis and application thereof in degradation of bletilla striata polysaccharide |
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| 平文祥等.沤麻系统中一株甘露聚糖酶产生菌产酶条件的优化.《黑龙江大学自然科学学报》.2008,第25卷(第6期), * |
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