CN1844366A - Halobacterium jilantaiense and application thereof - Google Patents

Abstract

This invention discloses a strain of Halobacterium jilantaiense and the application thereof. This strain is Halobacterium jilantaiense AC3 CGMCC No.1594, whose 16S rRNA encoding gene has the nucleotide sequence indicated by the No.1 sequence in the sequence table. The strain can produce lipase with tolerance to high salt concentration. The auxocyte, cell suspension, or the lipasse can catalyse the hydrolysis and synthesis of lipids in the range of 1.0-5.0M concentration of NaCl. And the lipase produced by this strain can catalyse triglyceride and other glycerides in high salt concentration solution or organic solvent.

Description

A Strain of Halobacterium Jilantai and Its Application

technical field

The invention relates to a strain of halophilic bacillus gilantai and its application, in particular to the application of the strain of halophilic bacillus gilantai in the production of lipase.

Background technique

Lipase (lipase, EC 3.1.1.3) is widely distributed in nature and catalyzes the synthesis of triglyceride from fatty acid and glycerol and its reverse reaction. Lipase is a very important industrial enzyme, which is widely used in the processing and manufacturing of food, cosmetics, leather, paper, and washing products. Lipase catalyzes the hydrolysis and synthesis of water-insoluble triglycerides. The catalytic reaction is heterogeneous catalysis at the oil-water interface. The reaction rate is not only limited by the catalytic ability of the enzyme itself, but also depends on the size of the contact interface between the substrate and the enzyme. related.

In many fields of application of lipase, catalytic conditions with high salt concentration often appear, such as the field of environmental protection or the field of food processing. The activity of ordinary lipase is reduced or completely inactivated under the condition of high salt concentration, which limits its application.

There is currently a lack of lipases that can tolerate high salt concentrations. Extreme halophilic archaea are an important branch of extremophile research, widely distributed in some natural or artificial high-salt environments such as salt lakes, salt mines, salt ponds and preserved foods. The demand for NaCl of extreme halophilic archaea is not less than 1.5M, and the optimum growth salt concentration is generally 3.0-4.5M. In order to resist the osmotic pressure generated by the high salt concentration outside the cell, the intracellular accumulation of KCl at a concentration close to saturation . During long-term evolution, enzymes produced by extreme halophilic archaea are quite stable in high concentrations of KCl or NaCl solutions. Compared with ordinary proteins, extreme halophilic enzymes have a higher proportion of acidic amino acid residues, and the proportion of hydrophobic amino acids is significantly reduced, which reflects that the hydrophobic interaction is enhanced under the condition of high salt concentration, and reducing the proportion of hydrophobic amino acids is beneficial Prevent the protein from being overly compressed, thereby maintaining the activity and solubility of the protein under the condition of high salt concentration. Similar to organic solvents, salts in solution are also effective in reducing the activity of water. For example, the activity of water in saturated NaCl solution is 0.75, which is equivalent to the activity of water in 60% N,N-dimethylformamide.

Contents of the invention

One object of the present invention is to provide a strain of Halobacterium gilantae capable of producing high-salt lipase.

The Halobacterium jilantaiense that can produce high-salt lipase is Halobacterium jilantaiense AC3, which was deposited in the "Common Microbiology Center of China Committee for Microbial Culture Collection" in Beijing on January 23, 2006 , and its deposit number is CGMCC No.1594.

Halobacterium jilantaiense AC3CGMCC No.1594 was isolated from the saline sediments of Jilantai Salt Lake (located in Jilantai Town, Alxa Left Banner, 102 kilometers north of Bayanhot Town).

Halobacterium jilantaiense (Halobacterium jilantaiense) AC3CGMCC No.1594, its 16SrDNA has the nucleotide sequence in sequence 1 in the sequence listing. The growth of the strain requires no less than 1.5M NaCl concentration, can grow in the range of 2.5-5.0mol/L salt concentration, and the optimum NaCl concentration is 3.0-4.5M. This strain can produce lipase resistant to high concentration of salt. The grown cells or cell suspension or the prepared lipase can catalyze the hydrolysis and synthesis of lipid substances in the range of NaCl concentration 1.0-5.0M, such as the hydrolysis of Tween series compounds and synthesis; in addition, similar to organic solvents, the salt in the solution also has the effect of effectively reducing the activity of water, the strain and its lipase also have activity and stability in organic solvents, and can be used for lipids in organic solvents Catalytic reactions for the hydrolysis and synthesis of similar substances.

Another object of the present invention is to provide a lipase that can catalyze the hydrolysis and synthesis of lipids under high-salt conditions.

The lipase provided by the invention is a metabolite obtained by fermenting Halobacterium jilantaiense AC3CGMCC No.1594.

The Halobacterium jilantaiense AC3 CGMCC No.1594 can be used in the inorganic salt medium with sugar alcohol compound as the only carbon source and energy source. Grow and produce lipase, such as CM medium (casamino acid (Casamino acid, Difco) 7.5g, yeast extract (Oxoid) 10.0g, MgSO ₄ 7H ₂ O 20.0g, trisodium citrate 3.0g, KCl 2.0g, FeSO ₄ 7H ₂ O 0.05g, NaCl 200g, distilled water 1000ml, adjust the pH to 7.0-7.4 with 1M NaOH, add 15g agar to the solid medium), milk salt medium (150mL skimmed milk, inorganic salt solution ( MgSO ₄ 7H ₂ O 3g, NaCl 60g, KNO ₃ 0.6g, 0.5mg ferric citrate) 30mL, agar solution (Casamino acid 1.5g, glycerin 3g, agar 4.5g) 120ml) or in the presence of some sugar alcohol derivatives , such as: alanine, pyruvic acid, potassium acetate, citrate, malonate, glucose, fructose, xylose, galactose, mannose, sucrose, maltose as the only carbon source inorganic salt basal medium (the The formula of inorganic salt basal medium is: MgSO ₄ 7H ₂ O 20.0g, trisodium citrate 3.0g, KCl 2.0g, FeSO ₄ 7H ₂ O 0.05g, NaCl 200g, distilled water 1000ml, adjust pH to 7.0-7.4, add 15g agar to the solid medium).

The above-mentioned lipase can catalyze the decomposition and synthesis reaction of lipid compounds under the condition of high salt ion concentration (such as Na ⁺ concentration is 1.0-5.0mol/L) and organic solvent phase, such as the saturated fatty acid carbon chain length of 4-18 And unsaturated triglycerides and their mixtures, such as glyceryl tributyrate, glyceryl stearate, glyceryl palmitate, olive oil, palm oil. Surfactants containing ester bonds such as Tween 20, Tween 40, Tween 60, Tween 80, and fatty acid 4-nitrophenyl ester compounds (butyric acid 4-nitrophenyl ester, palmitic acid 4-nitrophenyl Phenyl ester, 4-nitrophenyl laurate compound) can also be used as the catalytic substrate of the lipase.

In addition to the stability and activity under high salt concentration conditions, the lipase of the present invention also has activity and stability in organic solvents, can be used for catalytic reactions in organic solvents, and can be used under high salt ion concentration conditions and in organic solvents. Catalyze the hydrolysis and synthesis of triglyceride and other oil esters in organic solvent phase.

Description of drawings

Figure 1 is the growth curve of Halobacterium jilantaiense AC3 CGMCC No.1594 under different NaCl concentrations.

Figure 2 is the catalytic activity curve of high-salt lipase produced by Halobacterium jilantaiense AC3 CGMCC No.1594 under different NaCl concentrations.

Figure 3 is the catalytic activity curve of high-salt lipase produced by Halobacterium jilantaiense AC3C GMCC No.1594 at different temperatures.

Fig. 4 is the relationship curve between the high-salt lipase activity produced by Halobacterium jilantaiense AC3 CGMCC No.1594 and the percentage content of the added induction substrate olive oil.

Detailed ways

The methods in the examples are conventional methods unless otherwise specified.

Example 1, Isolation and Identification of Halobacterium jilantaiense AC3 CGMCC No.1594

1. The acquisition and biological characteristics of Halobacterium jilantaiense AC3 CGMCC No.1594

Show through culture experiment, this bacterium has the following characteristics: (1) colony characteristic: on milk salt plate (skim milk 150mL, inorganic salt solution (MgSO _{7H 2} O 3g, NaCl 60g, KNO ₃ 0.6g, 0.5mg lemon (Casamino acid) 30mL, agar solution (Casamino acid 1.5g, glycerol 3g, agar 4.5g) 120ml) for 7 days, the colony was round, with a diameter of 0.5-2.1mm, smooth surface, neat edges, raised, cinnamon red . (2) Halobacterium jilantaiense AC3 CGMCC No.1594 strain grows strictly aerobically; the suitable pH range for growth is 6-9, and the optimum growth pH range is 7.5-8.5; the growth temperature range is 35-45°C, The optimum temperature is 40°C; the cells are short rod-shaped; Gram staining is negative; the cell size is 0.5μm×1.5μm. (3) Halobacterium jilantaiense AC3 CGMCCNo.1594 can hydrolyze starch, is positive for catalase and oxidase, does not produce fluorescent pigment, hydrolyzes starch and casein, can liquefy gelatin, and can utilize potassium propionate, Pyruvate, potassium acetate, citrate, malonate, glucose, fructose, xylose, galactose, mannose, sucrose, maltose, lactose and sorbose cannot be utilized.

2. PCR amplification and sequence determination of the 16S rRNA gene of Halobacterium jilantaiense AC3 CGMCC No.1594

Halobacterium jilantaiense (Halobacterium jilantaiense) AC3 CGMCC No.1594 was inoculated in CM liquid medium, and the fermentation broth grown to the late logarithmic period was centrifuged (12000 rpm, 5 minutes) to remove the supernatant, and 20% Wash 2-3 times with NaCl solution; beat the cells with 1mL 20% NaCl solution, add 2mL 10% SDS, 8mL sterile water and proteinase K to a final concentration of 50-100μg/mL; incubate at 55°C for 1-2 hours for treatment ; Add an equal volume of Tris saturated phenol, shake gently for 5 minutes, centrifuge (12000 rpm, 5 minutes), use a wide-mouthed tip to absorb the supernatant, and then sequentially phenol-chloroform-isoamyl alcohol (25:24: 1) and chloroform-isoamyl alcohol (24:1, volume ratio) for one treatment. Precipitate with 2 times the volume of ice ethanol, soak in 70% ice ethanol solution for 5 minutes. After drying, it was dissolved in TE solution as a template. For the PCR reaction of 16S rRNA gene amplification, the forward primer is 5'-TTCCGGTTGATCCTGCC-3' and the reverse primer is 5'-AAGGAGGTGATCCAGCC-3', corresponding to 7-23 and 1425- of the 16S rRNA gene of Escherichia coli, respectively 1541 bases. The sequencing of PCR products was completed by Chinese national human genome center-SinoGenoMax Beijing, China, using ABI BigDye3.1 sequencing kit (Applied Biosystems) and DNA automatic sequencer (model ABI3730; Applied Biosystems). The PCR reaction system (50 μL) was: 5 μL of 10×buffer, 4 μL of 25 mmol/L MgCl ₂ , 1 μL of 10 mmol/L dNTPs, 1 μL of each 30 pmol/L primer, 38 μL of ddH ₂ O, and 0.4 μL (2.5U) of Taq DNase. The PCR reaction conditions were as follows: 4 minutes at 95°C, 1 minute at 95°C, 1 minute at 45°C, 1 minute at 72°C, 30 cycles; finally 10 minutes at 72°C, and stored at 4°C. The length of the 16S rRNA gene sequence is 1402bp, and the similarity with the sequence of the strain Halobacterium jilantaiense NG4 ^T rRNA gene (accession number DQ256409) is 99%. The nucleotide sequence of its 16SrRNA gene is shown as sequence 1 in the sequence listing.

3. The taxonomic status of Halobacterium jilantaiense AC3 CGMCC No.1594

Referring to the content of "Bergey's Manual of Systematic Bacteriology", 2 ^nd ed, vol1, pp301-305, according to its above-mentioned morphological characteristics, physiological and biochemical characteristics, and the search results of its 16S rRNA gene sequence in GenBank, the strain AC3 (CGMCC No.1594) is Halobacterium jilantaiense, whose Latin name is Halobacterium jilantaiense. The Halobacterium jilantaiense AC3 strain was deposited in Beijing on January 23, 2006 in the "General Microorganism Center of China Committee for Culture Collection of Microorganisms", and its preservation number is CGMCC No.1594.

Embodiment 2, the growth situation of Jilantai halophilic bacillus (Halobacterium jilantaiense) AC3 CGMCC No.1594 under the condition of different NaCl concentrations and the substrate scope of the lipase that produces

1. Growth of Halobacterium jilantaiense AC3 CGMCC No.1594 under different NaCl concentrations

Inoculate Halobacterium jilantaiense AC3 CGMCC No.1594 in 0.75g/100ml casamino acid (Casamino acid, Difco), 1g/100ml yeast extract (Yeast extract, Oxoid), 0.3 g/100ml (0.068M) trisodium citrate trihydrate, magnesium sulfate heptahydrate 2.0g/100ml (0.081M), potassium chloride 0.2g/100ml (0.0268M), ferrous sulfate heptahydrate 0.05g/100ml (0.00018 M), sodium chloride concentrations are 6g/100ml (1.03M), 8g/100ml (1.36M), 10g/100ml (1.7M), 12g/100ml (2.05M), 14g/100ml (2.40M), 16g /100ml(2.74M), 18g/100ml(3.08M), 20g/100ml(3.42M), 22g/100ml(3.76M), 24g/100ml(4.11M), 26g/100ml(4.45M), 28g/100ml (4.80M), 30g/100ml (5.13M) and 1000ml of distilled water in a liquid medium (adjust the pH to 7.0-7.4 with 1M NaOH), after culturing at 40°C for 3 days, observe the growth of the strain, and detect the absorbance of the bacterial solution. The results are shown in Figure 1, indicating that the strain can grow in the range of 2.5-5.0M salt concentration, and the optimum salt concentration is 3.0-4.5M.

2. The substrate range of lipase action of Halobacterium gilantae AC3 CGMCC No.1594

Halobacterium gilantae AC3 CGMCC No.1594 was inoculated in saturated or unsaturated triglycerides (tributyrin, stearin, glycerin palmitate) containing 1mmol/L carbon chain length of 4-18, respectively. Fat, olive oil, palm oil), 1.0g/100mL Tween 20, 1.0g/100mL Tween 40, 1.0g/100mL Tween 60, 1.0g/100mL Tween 80, or CM of 1.0g/100mL olive oil Solid medium, cultured under light at 37°C for 5-7 days, observe the transparent circle or halo around the colony, the results show that in saturated or unsaturated triglycerides with a carbon chain length of 4-18, olive oil, Tween 20, Tween Halos formed around the substrates such as Tween 40, Tween 60, and Tween 80, which indicated that they could act as substrates for the lipase. The activity of lipase was calculated by detecting the increase of p-nitrophenol concentration at 410nm due to the decomposition of p-nitrophenol palmitate (p-NPP) under the hydrolysis of lipase with a UV spectrophotometer. Concrete steps: to reaction system (0.05g/100ml p-NPP, 5mmol/L sodium acetate-acetic acid buffer solution of pH7.0, 1% (volume concentration) Triton X-100, 5% (volume concentration,) Virahol ) was added 1 mL enzyme solution (0.10 g lipase dissolved in 1 mL pH6.8 Tris-HCL buffer), and the reaction was started at 37° C. for 5 minutes, and the molar extinction coefficient of p-nitrophenol was 12.75 mmol ⁻¹ cm ⁻¹ . The lipase activity can be calculated according to the amount of p-nitrophenol produced per minute. Experiments show that the lipase also has a hydrolysis effect on fatty acid 4-nitrophenyl ester compounds such as 4-nitrophenyl butyrate, 4-nitrophenyl palmitate and 4-nitrophenyl laurate.

Embodiment 3, the mensuration of the lipase purification and enzyme activity that Jilantai halophilic bacillus (Halobacterium jilantaiense) AC3 CGMCC No.1594 produces:

1. Purification of lipase produced by Halobacterium jilantaiense AC3 CGMCC No.1594

Halobacterium jilantaiense AC3 CGMCC No.1594 was cultured in the following CM liquid medium: Casamino acid (Difco) 7.5g, Yeast extract (Oxoid) 10.0g, MgSO ₄ ·7H ₂ O 20.0g, Trisodium citrate 3.0g, KCl 2.0g, FeSO ₄ ·7H ₂ O 0.05g, NaCl 200g, distilled water 1000ml, adjust the pH to 7.0-7.4 with 1M NaOH, and sterilize at 121 degrees Celsius for 30 minutes. Cultivate at 40°C, light and ventilate (the amount of liquid filled is 1/5 of the volume of the shake flask, the radius of rotation is 35 mm, and 200 rpm) for 5-7 days. Centrifuge the cultured bacterial solution at 8,000-12,000 rpm for 5-10 minutes, add 20-30% volume of acetone to the supernatant, centrifuge at 8,000-12,000 rpm for 5-10 minutes, and store the collected precipitate on ice Dissolve in the pre-cooled 0.02mol/L Tris-HCl buffer solution (pH7.0) in the bath, and centrifuge at 8,000-12,000 rpm for 5-10 minutes to remove impurities and precipitates to obtain a crude enzyme solution. In an ice bath, slowly add finely ground ammonium sulfate into the crude enzyme solution while stirring, place at 4°C for 12 hours, centrifuge at 10,000 rpm for 15 minutes, and collect the protein precipitated by ammonium sulfate with a saturation of 20-75% , dissolved with 0.02mol/L Tris-HCl buffer (pH7.0) and dialyzed with deionized water to remove ammonium sulfate. DEAE-cellulose (DE52) ion exchange chromatography, equilibrated with 0.02mol/LTris-HCl buffer (pH7.0). Put the dialyzed enzyme solution on the column, the flow rate of the column is 30ml/hour, first elute with the initial buffer (0.01Mol/L pH7.4PB solution), and then use the gradient elution of 0-1.0mol/L sodium chloride solution , the flow rate is 30ml/hour, the activity peaks are combined and then concentrated. Equilibrate Sephadex G-100 with 0.02mol/L Tris-HCl buffer (pH7.0), purify and concentrate the enzyme solution through a DEAE-cellulose (DE52) ion exchange column, and carry out Sephadex G-100 chromatography, use 0.02 mol/L Tris-HCl buffer (pH7.0) was used for elution, and the flow rate of the eluent was 18ml/hour. After combining the enzyme activity peaks, the lipase powder was obtained after concentration and drying at 20°C. The results showed that under the experimental conditions, the output of lipase powder was 35mg/L culture medium.

2. Determination of enzyme activity

Accurately weigh 0.100 g of the lipase enzyme powder purified in step 1, add a small amount of water and mix thoroughly to form a paste, add water to make up to 10 mL, and make a 100-fold diluted enzyme solution for enzyme activity determination. The enzyme activity assay of lipase adopts following titration method and spectrophotometer method:

(1) Titration method

One enzyme activity unit of lipase is defined as the amount of enzyme that produces 1 μmol of fatty acid per minute when lipase hydrolyzes triglycerides at 37°C and pH 7.0.

The measurement principle is: when the reaction time of lipase activity measurement is short (3-15 minutes), it can be considered that the reaction catalyzed by lipase is to generate one molecule of fatty acid and one molecule of diglyceride per molecule of triglyceride. Using olive oil as a substrate, the fatty acid released by hydrolysis can be titrated with a standard alkaline solution. In the case of a complete excess of the substrate, the titration value is used to indicate the enzyme activity. The activity of lipase was determined according to the following method: Take a 100mL Erlenmeyer flask, add 5mL of 0.025mol/L phosphate buffer with a pH of 7.0 and 4mL of polyvinyl alcohol olive oil emulsion to each bottle, and put it in a water bath at 37°C (enzymolysis temperature) Preheat a 100mL Erlenmeyer flask in medium, then add 1mL of enzyme solution, accurately time the enzyme solution from the beginning, keep warm for 15 minutes, and immediately add 15mL of absolute ethanol to stop the enzyme reaction. Add three drops of phenolphthalein indicator, drop to reddish with 0.05mol/L sodium hydroxide standard solution, and make a blank control at the same time, add ethanol in the control sample before the enzyme solution, no need to keep warm. Three parallel runs were made for each sample, and the average value was taken. The results showed that the specific activity of the lipase powder purified in step 1 was 55U/mg.

(2) Spectrophotometer method

One unit (U) of enzyme activity is defined as the amount of enzyme required to convert 1 μmol of p-NPP per minute under the conditions of 37°C and pH 7.0.

The principle of determination is as follows: use a UV spectrophotometer to detect the increase of p-nitrophenol concentration at 410nm due to the decomposition of p-nitrophenol palmitate (p-NPP) under the hydrolysis of lipase, and calculate the activity of lipase. The reaction system includes 0.05% (0.05g/100ml) p-NPP, pH 7.0 5mmol/L sodium acetate-acetic acid buffer, 1% (volume concentration) Triton X-100, 5% (volume concentration) isopropanol; 50mM Tris-HCL buffer (pH6.8), 1mL enzyme solution. The molar extinction coefficient of p-nitrophenol = 12.75 mmol ^-1 cm ^-1 . The results showed that the specific activity of the lipase powder purified in step 1 was 54.3U/mg.

The titration method is the main method, and the spectrophotometer method is used as the verification method. The results show that the lipase activity values measured by the two methods are very close

Embodiment 4, the characteristics of the lipase produced by Halobacterium jilantaiense AC3 CGMCC No.1594

1. Catalytic activity under different concentrations of NaCl

The lipase powder purified in embodiment 3 step 1 adopts the spectrophotometer method in embodiment 3 step 2 to contain 0mol/L, 0.5mol/L, 1.0mol/L, 1.5mol/L, 2.0mol/L, The enzyme activity was measured in the reaction system of 2.5mol/L, 3.0mol/L, 3.5mol/L, 4.0mol/L, 4.5mol/L, 5.0mol/L NaCl, the results are shown in Figure 2, indicating that Jilantai The lipase produced by Halobacterium jilantaiense AC3 CGMCCNo.1594 has catalytic activity in the NaCl range of 1.0mol/L-5.0mol/L, and the optimum salt concentration range is 3.0mol/L-5.0mol/L.

2. Catalytic activity under different temperature conditions

The lipase powder purified in Step 1 of Example 3 was tested at 28°C, 30.6°C, 32.1°C, 32.5°C, 33.6°C, 34.3°C, 35°C, 36°C, 37°C using the spectrophotometer method in Step 2 of Example 3 ℃, 38.5 ℃, 40 ℃, 42 ℃, 45 ℃, and 50 ℃, the enzyme activity was measured, and the results are shown in Figure 3, indicating that the lipase produced by Halobacterium jilantaiense (Halobacterium jilantaiense) AC3 CGMCC No.1594 It has catalytic activity in the range of 28°C-50°C, and the optimum temperature range is 37°C-42°C.

Embodiment 5, the hydrolysis of olive oil by Halobacterium jilantaiense AC3 CGMCC No.1594 in CM liquid medium

The halophilic bacteria AC3 CGMCC No.1594 strain culture solution of Gilanthai halophilus AC3 CGMCC No.1594 cultured in the CM liquid medium to OD ₆₀₀ =0.2 was inoculated in the addition volume percentage concentration of 0% by 1% (volume ratio) , 0.50%, 1.00%, 1.50%, 2.00%, 2.50%, 3.00%, 4.00%, 5.00%, 6.00% olive oil in the CM liquid medium, at 37 ° C, 180r/min shaker culture, 72 Sampling and measuring thalline growth in hours, adopting the spectrophotometer method in the step 2 of embodiment 3 to measure the relationship between the lipase enzyme activity in the fermented liquid and the amount of olive oil added. The results show that olive oil plays a significant role in inducing lipase, and the results of lipase activity under different olive oil concentrations are shown in Figure 4. higher.

Sequence Listing

<160>1

<210>1

<211>1402

<212>DNA

<213> Halobacterium jilantaiense

<400>1

atcgatttag ccatgctagt tgtgcgggtt tagacccgca gcggaaagct cagtaacacg 60

tggccaagct accctgtgga cgggaataat ctcgggaaac tgaggctaat tcccaataac 120

gcttcacccc tggaatgggt gaagccggaa acgttccgac gccacaggat gcggctgcgg 180

tcgattaggt agacggtggg gtaacggccc accgtgccta taatcggtac gggttgtgag 240

agcaagagcc cggagacgga atctgagaca agattccggg ccctacgggg cgcagcaggc 300

gcgaaacctt tacactgtac gcaagtgcga taaggggact ccgagtgtga aggcatagag 360

ccttcacttt tgtacaccgt aaggtggtgt acgaataagg actgggcaag accggtgcca 420

gccgccgcgg taataccggc agtccgagtg atggccgata ttatgggcc taaagcgtcc 480

gtagctggct agacaagtcc gttgggaaat ctgcccgctt aacgggcagg cgtccagcgg 540

aaactgtcta gcttgggacc ggaagacctg aggggtacgt ctggggtagg agtgaaatcc 600

tgtaatcctg gacggaccgc cggtggcgaa agcgcctcag gagaacggat ccgacagtga 660

gggacgaaag ctagggtctc gaaccggatt agatacccgg gtagtcctag ctgtaaacga 720

tgtccgctag gtggtggcgc aggctacgag cctgcgctgt gccgtaggga agccgagaag 780

cggaccgcct gggaagtacg tctgcaagga tgaaacttaa aggaattggc gggggagcac 840

tacaaccgga ggagcctgcg gtttaattgg actcaacgcc ggacatctca ccggtcccga 900

cagtagtaat gacggtcagg ttgacgactt tacccgacgc tactgagagg aggtgcatgg 960

ccgccgtcag ctcgtaccgt gaggcgtcct gttaagtcag gcaacgagcg agacccacac 1020

ttctagttgc cagcaacacc cctgcggtgg ttgggtacac taggaggact gccattgcta 1080

aaatggagga aggaatgggc aacggtaggt cagtatgccc cgaatggacc gggcaacacg 1140

cgggctacaa tggctatgac agtgggatgc aacgccgaga ggcgacgcta atctccaaac 1200

gtagtcgtag ttcggattgc gggctgaaac ccgcccgcat gaagctggat tcggtagtaa 1260

tcgcgtgtca gaagcgcgcg gtgaatacgt ccctgctcct tgcacacacc gcccgtcaaa 1320

gcacccgagt gggtccgga tgaggccgtc atgcgacggt cgaatctggg ctccgcaagg 1380

gggcttaagt cgtaacaaga gc 1402

Claims (10)

1. Halobacterium jilantaiense AC3 CGMCC No.1594. 2. Halobacterium jilantaiense AC3 CGMCC No.1594 according to claim 1, characterized in that: the 16S rDNA of Halobacterium jilantaiense AC3 CGMCC No.1594 has the sequence Nucleotide sequence in sequence 1 in the list. 3. A lipase obtained by fermenting Halobacterium jilantaiense AC3CGMCC No.1594. 4. The lipase according to claim 3, characterized in that: the fermentation medium of the Halobacterium jilantaiense AC3 CGMCC No.1594 is composed of alanine, pyruvic acid, potassium acetate, lemon salt, malonate, glucose, fructose, xylose, galactose, mannose, sucrose or maltose as the only carbon source inorganic salt basal medium, CM medium or milk salt medium; The composition of the CM medium is 7.5g of Casamino acid, 10.0g of yeast extract, 0.0g of MgSO ₄ ·7H ₂ O2, 3.0g of trisodium citrate, 2.0g of KCl, 0.05g of FeSO ₄ ·7H ₂ O, NaCl 200g, 1000ml distilled water, pH7.5-8.5, add 15g agar to the solid medium; The composition of described milk salt medium is skimmed milk 150mL, inorganic salt solution 30mL, the aqueous solution 120ml that contains 1.5gCasamino acid and 3g glycerol, adds 4.5g agar in the solid medium, pH7.5-8.5; The described inorganic salt solution The composition is MgSO ₄ ·7H ₂ O 3g, NaCl 60g, KNO ₃ 0.6g, 0.5mg ferric citrate, 30mL water. 5. The lipase according to claim 4, characterized in that: the composition of the inorganic salt basal medium is: MgSO ₄ ·7H ₂ O 20.0g, trisodium citrate 3.0g, KCl 2.0g, FeSO ₄ · 7H ₂ O 0.05g, NaCl 200g, distilled water 1000ml, pH7.5-8.5, add 15g agar to the solid medium. 6. The application of the lipase according to claim 3, 4 or 5 in the decomposition or synthesis of lipid compounds. 7. The application according to claim 6, characterized in that the lipase catalyzes the decomposition or synthesis of lipid compounds under high-salt conditions. 8. The application according to claim 7, characterized in that: the high-salt condition is that the Na ⁺ concentration is 1.0-5.0 mol/L. 9. The application according to claim 6, 7 or 8, characterized in that: the lipid compound is triglyceride, Tween 20, Tween 40, Tween 60, Tween 80 or fatty acid 4-nitro Phenyl esters. 10. The application according to claim 9, characterized in that: the triglyceride is glyceryl tributyrate, glyceryl stearate, glyceryl palmitate, olive oil or palm oil; the fatty acid 4- The nitrophenyl ester compound is 4-nitrophenyl butyrate, 4-nitrophenyl palmitate or 4-nitrophenyl laurate.

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