CN116496942A - Strain for producing D-tagatose and application thereof - Google Patents
Strain for producing D-tagatose and application thereof Download PDFInfo
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- LKDRXBCSQODPBY-OEXCPVAWSA-N D-tagatose Chemical compound OCC1(O)OC[C@@H](O)[C@H](O)[C@@H]1O LKDRXBCSQODPBY-OEXCPVAWSA-N 0.000 title claims abstract description 59
- 238000006243 chemical reaction Methods 0.000 claims abstract description 39
- FBPFZTCFMRRESA-GUCUJZIJSA-N galactitol Chemical compound OC[C@H](O)[C@@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-GUCUJZIJSA-N 0.000 claims abstract description 20
- 239000000758 substrate Substances 0.000 claims abstract description 18
- 230000000284 resting effect Effects 0.000 claims abstract description 12
- 238000000034 method Methods 0.000 claims abstract description 8
- 230000003197 catalytic effect Effects 0.000 claims abstract description 4
- 238000000855 fermentation Methods 0.000 claims description 10
- 230000004151 fermentation Effects 0.000 claims description 10
- 239000000243 solution Substances 0.000 claims description 10
- 239000007853 buffer solution Substances 0.000 claims description 7
- 238000005119 centrifugation Methods 0.000 claims description 6
- 239000007788 liquid Substances 0.000 claims description 6
- 239000000872 buffer Substances 0.000 claims description 4
- 238000011218 seed culture Methods 0.000 claims description 4
- 238000005406 washing Methods 0.000 claims description 4
- 238000012262 fermentative production Methods 0.000 claims description 2
- 239000013049 sediment Substances 0.000 claims description 2
- 235000013305 food Nutrition 0.000 abstract description 4
- 102000004316 Oxidoreductases Human genes 0.000 abstract description 3
- 108090000854 Oxidoreductases Proteins 0.000 abstract description 3
- 238000003786 synthesis reaction Methods 0.000 abstract description 3
- 238000009412 basement excavation Methods 0.000 abstract description 2
- 230000015572 biosynthetic process Effects 0.000 abstract description 2
- 230000000694 effects Effects 0.000 description 12
- 239000002609 medium Substances 0.000 description 11
- 241000193830 Bacillus <bacterium> Species 0.000 description 7
- LONLXRPIYFRSMN-WNQIDUERSA-N (2r)-2-amino-3-sulfanylpropanoic acid;9h-carbazole Chemical compound SC[C@H](N)C(O)=O.C1=CC=C2C3=CC=CC=C3NC2=C1 LONLXRPIYFRSMN-WNQIDUERSA-N 0.000 description 6
- 239000000047 product Substances 0.000 description 6
- 239000006228 supernatant Substances 0.000 description 6
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 5
- 239000001963 growth medium Substances 0.000 description 5
- 239000000203 mixture Substances 0.000 description 5
- 239000000843 powder Substances 0.000 description 5
- WQZGKKKJIJFFOK-SVZMEOIVSA-N (+)-Galactose Chemical compound OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@H]1O WQZGKKKJIJFFOK-SVZMEOIVSA-N 0.000 description 4
- UJOBWOGCFQCDNV-UHFFFAOYSA-N 9H-carbazole Chemical compound C1=CC=C2C3=CC=CC=C3NC2=C1 UJOBWOGCFQCDNV-UHFFFAOYSA-N 0.000 description 4
- 241001052560 Thallis Species 0.000 description 4
- 238000009472 formulation Methods 0.000 description 4
- 238000012216 screening Methods 0.000 description 4
- 238000000926 separation method Methods 0.000 description 4
- 239000002689 soil Substances 0.000 description 4
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- 239000001888 Peptone Substances 0.000 description 3
- 108010080698 Peptones Proteins 0.000 description 3
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 3
- 239000012084 conversion product Substances 0.000 description 3
- 235000019319 peptone Nutrition 0.000 description 3
- 238000004321 preservation Methods 0.000 description 3
- 229920001817 Agar Polymers 0.000 description 2
- 108010018080 L-arabinose isomerase Proteins 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- 239000008272 agar Substances 0.000 description 2
- 239000003513 alkali Substances 0.000 description 2
- 230000001580 bacterial effect Effects 0.000 description 2
- 230000000975 bioactive effect Effects 0.000 description 2
- 238000011088 calibration curve Methods 0.000 description 2
- 238000006555 catalytic reaction Methods 0.000 description 2
- 238000012258 culturing Methods 0.000 description 2
- 230000007423 decrease Effects 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 229940079593 drug Drugs 0.000 description 2
- 238000002955 isolation Methods 0.000 description 2
- 229910021645 metal ion Inorganic materials 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 239000012086 standard solution Substances 0.000 description 2
- 230000009466 transformation Effects 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 108020004465 16S ribosomal RNA Proteins 0.000 description 1
- 244000063299 Bacillus subtilis Species 0.000 description 1
- 235000014469 Bacillus subtilis Nutrition 0.000 description 1
- 102000015833 Cystatin Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 102000002464 Galactosidases Human genes 0.000 description 1
- 108010093031 Galactosidases Proteins 0.000 description 1
- 108090000769 Isomerases Proteins 0.000 description 1
- 102000004195 Isomerases Human genes 0.000 description 1
- 239000012880 LB liquid culture medium Substances 0.000 description 1
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 1
- VLSOAXRVHARBEQ-UHFFFAOYSA-N [4-fluoro-2-(hydroxymethyl)phenyl]methanol Chemical compound OCC1=CC=C(F)C=C1CO VLSOAXRVHARBEQ-UHFFFAOYSA-N 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 230000003178 anti-diabetic effect Effects 0.000 description 1
- PYMYPHUHKUWMLA-UHFFFAOYSA-N arabinose Natural products OCC(O)C(O)C(O)C=O PYMYPHUHKUWMLA-UHFFFAOYSA-N 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- SRBFZHDQGSBBOR-UHFFFAOYSA-N beta-D-Pyranose-Lyxose Natural products OC1COC(O)C(O)C1O SRBFZHDQGSBBOR-UHFFFAOYSA-N 0.000 description 1
- 239000012620 biological material Substances 0.000 description 1
- 239000012295 chemical reaction liquid Substances 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
- 238000000576 coating method Methods 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 239000002537 cosmetic Substances 0.000 description 1
- 108050004038 cystatin Proteins 0.000 description 1
- 230000002950 deficient Effects 0.000 description 1
- -1 diluting Substances 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 235000003599 food sweetener Nutrition 0.000 description 1
- 239000002054 inoculum Substances 0.000 description 1
- 238000006317 isomerization reaction Methods 0.000 description 1
- BJHIKXHVCXFQLS-PQLUHFTBSA-N keto-D-tagatose Chemical compound OC[C@@H](O)[C@H](O)[C@H](O)C(=O)CO BJHIKXHVCXFQLS-PQLUHFTBSA-N 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 230000031700 light absorption Effects 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 230000002906 microbiologic effect Effects 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000011392 neighbor-joining method Methods 0.000 description 1
- 239000002417 nutraceutical Substances 0.000 description 1
- 235000021436 nutraceutical agent Nutrition 0.000 description 1
- 230000003334 potential effect Effects 0.000 description 1
- 235000013406 prebiotics Nutrition 0.000 description 1
- 230000035484 reaction time Effects 0.000 description 1
- 238000006479 redox reaction Methods 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 239000008223 sterile water Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 239000003765 sweetening agent Substances 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/02—Monosaccharides
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
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- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Genetics & Genomics (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Biomedical Technology (AREA)
- Virology (AREA)
- Tropical Medicine & Parasitology (AREA)
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Abstract
The invention discloses a strain for producing D-tagatose and application thereof, wherein the strain is classified and named as endophytic bacillusPriestia endophytica) Z723 accession number GDMCC No: 63266D-tagatose is produced by whole cell catalytic conversion of a substrate using resting cells of the strain. The strain utilizes galactitol as a substrate to produce D-tagatose through a whole-cell catalytic method, the conversion rate can reach 92%, a novel strain is provided for high-conversion-rate D-tagatose synthesis and related oxidoreductase excavation, and the strain has wide application prospects in the fields of food and medical treatment.
Description
Technical Field
The invention belongs to the technical field of microorganisms, and particularly relates to a strain for producing D-tagatose and application thereof.
Background
Currently, consumer preferences have undergone a paradigm shift from synthetic compounds to natural bioactive molecules, as these natural bioactive molecules are considered safe without any side effects. Among potential nutraceuticals, D-tagatose is of interest because of its low caloric value, prebiotic potential and antidiabetic properties, and the possibility to prevent lifestyle-related diseases, but D-tagatose is only present in small amounts in nature. D-tagatose, which is a novel sweetener, has low calorie, low absorption and other functional health care effects, meets the requirements of people on sweetness and health, and is widely applied to the fields of foods, medicines, cosmetics and other fields.
The most commonly reported route to D-tagatose is to isomerize D-galactose by L-arabinose isomerase. For example, rao et al constructed a genetically engineered Bacillus subtilis strain, converted lactose using arabinose isomerase and galactosidase, and added 3 mM Mn 2+ With whole cell catalysis, tagatose conversion was only 57.2%. D-galactose and D-tagatose exist in the reaction solution at the same time, so that the yield is low, and the downstream product separation is not facilitated. Moreover, most L-arabinose isomerase showed strict metal ion dependence on D-tagatose production (e.g.Mn 2+ ) Affecting the application of D-tagatose in foods and medicines. The redox reaction allows the almost complete conversion of galactitol to D-tagatose, and is free of metal ion dependence, with the potential to synthesize D-tagatose by alternative isomerization routes, but the only microorganisms reported so far that catalyze galactitol to produce D-tagatose areMycobacterium smegmatisThese few species and the related oxidoreductases are relatively deficient.
Disclosure of Invention
The first object of the present invention is to provide a novel D-tagatose-producing strain classified and named as endophytic bacillusPriestia endophytica) Z723 accession number GDMCC No:63266.
a second object of the present invention is to provide the use of the above strain in the fermentative production of D-tagatose.
As a preferred embodiment, D-tagatose is produced by whole cell catalytic conversion of a substrate using resting cells of the strain.
As a preferred embodiment, after seed culture is carried out on the strain, seed solution is inoculated to a fermentation medium for fermentation culture, fermentation liquid is taken for centrifugation to collect thalli, the thalli are subjected to resuspension washing in PBS buffer solution, and sediment, namely resting cells, are taken after centrifugation again.
As a preferred embodiment, the resting cells of the strain are resuspended in PBS buffer containing substrate and reacted with shaking to prepare D-tagatose.
As a preferred embodiment, the medium for seed culture is LB liquid medium.
As a preferred embodiment, the strain produces D-tagatose using galactitol as a substrate.
As a preferred embodiment, the substrate concentration in the conversion system is from 10 to 60g/L.
As a preferred embodiment, the pH in the conversion system is from 6 to 10; preferably 7-9.
As a preferred embodiment, the temperature in the conversion system is from 25 to 50℃and preferably from 40 to 50 ℃.
As a preferred embodiment, the reaction time of the conversion system is 12-24 hours.
The invention discloses a strain for preparing D-tagatose biologically, which is obtained by separating and screening from the soil of Xinjiang saline-alkali soil in China, and belongs to endophytic bacillus. The D-tagatose produced by utilizing galactitol has the conversion rate reaching 92%, provides a new strain for the synthesis of the D-tagatose with high conversion rate and the excavation of related oxidoreductase, and has wide application prospect in the fields of food and medical treatment.
Drawings
FIG. 1 shows the isolation and screening of D-tagatose-producing strains.
FIG. 2 is a view of the D-tagatose-producing strain observed under a microscope.
FIG. 3 shows a Z723 phylogenetic tree of endophytic Bacillus.
FIG. 4 is a calibration curve of the quantitative product D-tagatose by the cysteine carbazole chromogenic method.
FIG. 5 shows the growth curve of endophytic Bacillus Z723.
FIG. 6 shows the effect of pH of the reaction system on D-tagatose conversion.
FIG. 7 shows the effect of reaction temperature on D-tagatose conversion.
FIG. 8 is a graph showing the effect of galactitol substrate concentration on the conversion of D-tagatose.
FIG. 9 is a graph showing the conversion rate of D-tagatose by whole cell catalysis of galactitol by Bacillus endogenously Z723.
The biological material is classified and named as endophytic bacillusPriestia endophytica) Z723, deposited with the Guangdong province microbiological bacterial collection center (GDMCC), accession number GDMCC No:63266, date of preservation: 2023, 3, 12, deposit address: guangdong China.
Description of the embodiments
EXAMPLE 1 isolation screening and molecular biological identification of D-tagatose-producing Strain
1. Separation and screening of D-tagatose-producing strains
1. 5 g soil samples from Xinjiang saline-alkali soil in China are taken, diluted in 50 mL sterile water, filtered to remove sample residues, bacterial suspension is prepared, and transferred to 50 mL LB liquid medium (250 mL shake flask) for shake culture at 37 ℃ and 200 rpm for 24 h.
LB liquid medium formulation (g/L): peptone 10, naCl 10, yeast powder 5, pH 7.0.
2. Transferring the culture solution 1 mL to a fresh enrichment medium after the step 1 is completed, culturing at 200 rpm at 37 ℃ for 24 h to obtain the culture solution, diluting, coating 100 mu L of the culture solution on a plate of a separation medium, culturing at 37 ℃, after single colony grows on the plate, picking up a single colony, streaking and purifying three times, and inoculating to a slant culture medium for preservation.
Enrichment medium formulation (g/L): galactitol10. Yeast powder 5, (NH) 4 ) 2 SO 4 2.5、MgSO 4 7H 2 O 0.3、KH 2 PO 4 2、K 2 HPO 4 4，pH 7.0。
Plate separation media formulation (g/L): galactitol 10, yeast powder 5, (NH) 4 ) 2 SO 4 2.5、MgSO 4 7H 2 O 0.3、KH 2 PO 4 2、K 2 HPO 4 4, agar 20, pH 7.0.
Slant solid medium formula (g/L): galactitol 10, yeast powder 5, peptone 5, naCl 5, agar 20, pH 7.0.
3. After the step 2 is completed, the strain preserved on the slant culture medium is selected and inoculated into a fermentation culture medium, the strain is subjected to shaking culture at 37 ℃ and 200 rpm for 24 h, then the strain is collected by centrifugation, the strain is subjected to resuspension washing twice by using PBS buffer solution, supernatant is discarded after centrifugation again, resting cells are obtained, a certain amount of strain is subjected to shaking conversion 12 h by suspending in 10 mL of PBS buffer solution containing 20 g/L galactitol, and the strain is subjected to centrifugation for 10 min to obtain supernatant. Strains with bluish purple color change are screened by using a cysteine carbazole chromogenic method, as shown in figure 1, the reading is measured by using a spectrophotometer, the yield is calculated according to a D-tagatose standard curve, and one strain with the highest yield and highest enzyme activity is selected for further strain identification and is preserved at-80 ℃ by using glycerol.
Fermentation medium formulation (g/L): galactitol 20, peptone 10, yeast powder 5, naCl 10, KH 2 PO 4 2、K 2 HPO 4 4、MgSO 4 7H 2 O 0.3、CaCl 2 2H 2 O 0.02、MnSO 4 H 2 O 0.05、FeSO 4 7H 2 O 0.04, pH 7.0。
2. Molecular biological identification of D-tagatose-producing strains
The morphology of the strain was observed under a microscope, and the strain was in the form of a rod as shown in FIG. 2. The 16S rRNA sequence sequencing result of the strain is compared with the existing other preservation sequences through the online program NCBI by using BLAST, the comparison result is used for drawing a phylogenetic tree by using a Neighbor-Joining method in MEGA software, the result is shown in FIG. 3, and the strain is determined to be endophyticBacillus [ ]Priestia endophytica) This strain was designated as Z723.
Example 2 calibration curve of quantitative product D-tagatose by cystatin carbazole chromogenic method
1 mL of 1-50 mug/mL of D-tagatose standard solution is respectively added into 8 test tubes of 15 mL, 0.2 mL of 15 g/L L-cysteine hydrochloride solution, 6 mL of 70% sulfuric acid solution and 0.2 mL of 1.2 g/L carbazole alcohol solution are respectively added in sequence in the following order, after shaking and mixing, the mixture is cooled at room temperature for 10 min in a constant-temperature water bath of 60 ℃, the absorption value is measured at 550 nm by using a spectrophotometer, and the color development reaction liquid of D-tagatose is replaced by the added galactitol standard liquid to serve as a blank. And drawing a D-tagatose standard curve by taking the concentration of the D-tagatose standard solution as an abscissa and the light absorption value as an ordinate, and calculating the D-tagatose content through the standard curve as shown in figure 4.
EXAMPLE 3 determination of the growth curve of Bacillus Agrobacter Z723
Inoculating endophytic bacillus Z723 for producing D-tagatose into 5 mL LB liquid culture medium, placing at 37 ℃ and activating at 200 rpm for 12 h, and obtaining the product according to the inoculum size of 2%v/v) The fermentation medium was inoculated and incubated at 37℃and 200 rpm, and the absorbance of the culture medium was measured at each time point by sampling at intervals of 2 h, the total growth time was 24. 24 h, and the growth curve was plotted as shown in FIG. 5.
Example 4 influence of pH of reaction System, reaction temperature and concentration of galactitol substrate added in reaction System on D-tagatose conversion Effect
1. Influence of pH of reaction system on D-tagatose conversion effect
Inoculating endophytic bacillus Z723 for producing D-tagatose into a fermentation culture medium, performing shaking culture at 37 ℃ and 200 rpm for 24 h, centrifuging to collect thalli, re-suspending and washing twice by using PBS buffer solution, centrifuging again, discarding supernatant to obtain resting cells, suspending a certain amount of thalli in 1 mL PBS buffer solution containing 20 g/L galactitol and having pH value of 3.0-10.0, performing shaking reaction at 37 ℃ for 2 h, and centrifuging for 10 min to obtain supernatant. The concentration of the converted product D-tagatose was determined by cysteine carbazole chromogenic method at different pH systems to determine the optimal pH. As shown in FIG. 6, the optimum pH was 8.0.
2. Effect of reaction temperature on D-tagatose conversion Effect
The resting cells are resuspended in 1 mL PBS buffer solution containing 20 g/L galactitol and having a pH of 8.0, subjected to constant temperature shaking transformation at 25-50 ℃ for 2 h, and centrifuged for 10 min to obtain a supernatant. The concentration of D-tagatose as the conversion product at different reaction temperatures was determined by cysteine carbazole chromogenic method to determine the optimal reaction temperature. As shown in FIG. 7, 45℃is the optimum reaction temperature, and the yield tends to decrease at temperatures higher or lower than 45 ℃.
3. Effect of galactitol substrate concentration on D-tagatose conversion Effect
The resting cells were resuspended in 1, mL, PBS buffer, pH 8.0 containing 10, 20, 30, 40, 50 and 60g/L galactitol, and transformed with shaking at 37℃for 2 h, and centrifuged for 10 min to obtain the supernatant. The concentration of the conversion product D-tagatose at different galactitol substrate concentrations was determined using a cysteine carbazole chromogenic method to determine the optimal substrate concentration. As shown in FIG. 8, 20 g/L is the optimal substrate concentration, and the D-tagatose product concentration slightly increases when the substrate concentration is higher than 20 g/L, so that the cell and substrate combination tends to saturate, and the conversion rate decreases.
Example 5 optimal transformation conditionsPriestia endophyticaSynthesis of D-tagatose from Z723
Will bePriestia endophyticaZ723 resting cells were transformed with PBS buffer at pH 8.0 containing 20 g/L galactitol under constant temperature shaking at 45℃to give 24. 24 h% conversion, and the concentration of D-tagatose, a conversion product, was measured by cysteine carbazole chromogenic assay for different time periods, as shown in FIG. 9, at a conversion of 18.4 g/L D-tagatose after 24 h, at 92%.
Claims (10)
1. A strain for producing D-tagatose is characterized by being classified and named as endophytic bacillusPriestia endophytica) Z723 accession number GDMCC No:63266.
2. use of the strain of claim 1 for the fermentative production of D-tagatose.
3. The use according to claim 2, characterized in that D-tagatose is produced by whole cell catalytic conversion of the substrate with resting cells of the strain.
4. The method according to claim 3, wherein after the strain is subjected to seed culture, the seed solution is inoculated into a fermentation medium for fermentation culture, the fermentation solution is centrifuged to collect the cells, the cells are subjected to resuspension washing in PBS buffer solution, and the sediment, i.e. resting cells, is obtained after centrifugation.
5. Use according to claim 2 or 3, characterized in that D-tagatose is prepared by shaking the resting cells of the strain in a substrate-containing PBS buffer.
6. The use according to claim 2, wherein the medium for seed culture is LB liquid medium.
7. The use according to claim 2, wherein the strain produces D-tagatose on galactitol as a substrate.
8. Use according to claim 3, characterized in that the substrate concentration in the conversion system is 10-60g/L.
9. The use according to claim 5, wherein the pH in the conversion system is 3-10; preferably 7-9.
10. The use according to claim 5, wherein the temperature in the conversion system is 25-50 ℃; preferably 40-50 ℃.
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