CN104845922A - Acinetobacter sp. OUC-Qa2 and application of acinetobacter sp. OUC-Qa2 to synthesis of phosphatidylserine - Google Patents
Acinetobacter sp. OUC-Qa2 and application of acinetobacter sp. OUC-Qa2 to synthesis of phosphatidylserine Download PDFInfo
- Publication number
- CN104845922A CN104845922A CN201510325298.8A CN201510325298A CN104845922A CN 104845922 A CN104845922 A CN 104845922A CN 201510325298 A CN201510325298 A CN 201510325298A CN 104845922 A CN104845922 A CN 104845922A
- Authority
- CN
- China
- Prior art keywords
- ouc
- acinetobacter
- synthesis
- phosphatidylserine
- acinetobacter calcoaceticus
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The invention provides acinetobacter sp. OUC-Qa2. The preservation number of the strain is CGMCC NO.10471. The acinetobacter sp. OUC-Qa2 is used for biologically catalyzing phosphatidylcholine (PC) to synthesize phosphatidylserine (PS). The acinetobacter sp. OUC-Qa2 has high base exchange activity. After the acinetobacter sp. OUC-Qa2 is used for biologically catalyzing PC to synthesize PS, nuclear magnetic resonance analysis shows that the conversion rate of the substrate PC is 47.8%, the selectivity of PS synthesis is 74% and the quantity of phosphatidic acid (PA) produced by hydrolysis is low (26%). The acinetobacter sp. OUC-Qa2 has a very good application prospect.
Description
Technical field
The invention belongs to screening and the applied technical field thereof of functional bacteria, be specifically related to a kind of acinetobacter calcoaceticus and the application in phosphatidylserine synthesis thereof.
Background technology
After Uauquelin in 1812 finds Yelkin TTS in human brain, people find again the trace of Yelkin TTS in succession from yolk and plant seed.Yelkin TTS has good physiological function as the important component part of cytolemma, and simultaneously it also has moisturizing, emulsification, the effect such as anti-oxidant.Enter 21 century phosphatide and attract the concern of countries in the world as the third-largest healthcare products and food emulsifier.Phosphatide is commonly called as Yelkin TTS, narrow sense be refer in particular to phosphatidylcholine (Phosphatidyleholin, PC), be the general name that a class contains phosphatidyl glycerol ester mixture in a broad sense.Phosphatide can be divided into by the difference according to the phosphatidyl contained by phosphatide: phosphatidylcholine (Phosphatidylcholine; PC), phosphatidylethanolamine (Phosphatidylethanolamin; PE), phosphatidylinositols (Phosphatidylinositol; PL), phosphatidic acid Serine (Phosphatidylserine, PS), phosphatidyl glycerol (Phosphatidylglycero; and sphingomyelin (Springmyehn, SM) PG).
In numerous phospholipid fraction, PS is uniquely can the phosphorus of state of regulating cell film key protein, cytolemma can be helped to keep tough, maintain mobility, the permeability of cytolemma, and the metabolism that can activate multiple enzyme is synthesized.In addition, the neural system of PS on biology has the transmission that vital role can affect chemical message in brain, and helps brain cell to store and reading data, is the important element maintaining brain normal memory power, reaction and healthy mood.Research report, PS can improve the elderly A Erci Mohs disease (Alzheimen ' s Disease), repairs brain damage and improves cognitive power, keeping tensions down pressure and the effect such as physical fatigue, Cure of depression.PS extensively exists in the middle of daily food, but content rareness can not meet human body requirements; And after entering 30 years old with advancing age, PS and the phospholipid composition of human body reduce gradually, human body needs every day and supplements the pure PS of 100-300mg, so the production development of PS is significant.
The production of current PS mainly contains extraction method and Enzyme optrode.Extraction method mainly extracts PS from vegetable lecithin, pluck, cerebral tissue, extraction process is complicated and need a large amount of organic reagents, because the similarity chemical extraction of structure of phospholipid and character can only obtain crude product, be difficult to the single phosphatide preparing highly purified multi-functional characteristic, the use in the field such as medicine, food can not be met.Biological enzyme due to its have widely substrate specificity, reaction conditions is gentle, by product is few, wide material sources and the advantage such as convenient, has become the method that PS preparation has potentiality to be exploited most.The method is simple to operate, and organic reagent usage quantity is few, and the easy extraction and isolation of the product of enzymatic clarification, can greatly reduce PS production cost.
Phospholipase D (Phospholipid D, EC 3,1.4.4, abbreviation PLD) key of ester can be become with organic bases (as choline, thanomin etc.) hydroxyl by the phosphoric acid in catalytic hydrolysis phospholipid molecule, i.e. 4 ester bonds, generate phosphatidic acid (Phosphatidic acid, PA) and organic bases; Meanwhile, to be also combined with phosphatide base by the various oxy-compound of Baseexchange catalytic reaction (as Serine) and to form new phosphatide (as PS).Therefore, PLD Baseexchange activity may be used for the preparation of the directional transformation of phosphatide, the synthesis of medicine and single phosphatide, rare phosphatide, has important commercial value.The microorganism of the product Phospholipase D reported mainly contains streptomycete (Streptomyces), coryneform bacteria (Corynebacterium), intestinal bacteria (Escherichia coli), Salmonella (Salmonella), pseudomonas (Pseudomonad), Vibrio harveyi (Vibrio harveyi), bacillus cereus (Bacillus cereus), achromobacter (Achromobacter), aspergillus oryzae (Aspergillus oryzae).But, the microorganism that current major part produces Phospholipase D generates too much hydrolysate PA after catalytic substrate PC, and the amount of synthesis modification phosphatide (as PS) is lower, thus limit the application of PLD in phospholipid modified process such as preparation single phosphatide and rare phosphatide etc.Therefore, be necessary to screen a kind of bacterial strain that can reduce PA resultant quantity.
Summary of the invention
The object of this invention is to provide a kind of acinetobacter calcoaceticus and the application in phosphatidylserine synthesis thereof, thus make up the deficiencies in the prior art.
First the present invention provides a kind of acinetobacter calcoaceticus (Acinetobacter sp.) OUC-Qa2, this bacterial strain was deposited in China Committee for Culture Collection of Microorganisms's common micro-organisms center (the China General Microbiological Culture Collection Center being positioned at No. 3 Institute of Microorganism, Academia Sinica in Yard 1, BeiChen xi Road, Chaoyang District, Beijing City on 01 30th, 2015, CGMCC), deposit number is CGMCC NO.10471.
Acinetobacter calcoaceticus of the present invention is used for biocatalysis phosphatidylcholine (PC) synthetic phospholipid acyl Serine (PS).
It is active that acinetobacter calcoaceticus of the present invention (Acinetobacter sp.) OUC-Qa2 has high Baseexchange, after applying its catalyze phospholipid phatidylcholine (PC) synthetic phospholipid acyl Serine (PS), find that substrate PC transformation efficiency is 47.8% by nuclear magnetic resonance spectroscopy, the selectivity of PS synthesis is 74%, and hydrolysis produces the amount low (26%) of PA, there is extraordinary application prospect.
Accompanying drawing explanation
Fig. 1: the screening lithograph of bacterial strain OUC-Qa2 of the present invention;
Fig. 2: the evolution tree graph of bacterial strain OUC-Qa2 of the present invention;
Fig. 3: bacterial strain OUC-Qa2 of the present invention is used for phosphatidylserine composite diagram.
Embodiment
Below in conjunction with example, the present invention is described further.But condition used in embodiment can be selected according to prior art.For the method for actual conditions unreceipted in embodiment, can conveniently conditional operation.
Embodiment 1 bacterial strain screening
(1) bacterial strain screening and cultivation
The fabaceous lecithin press residue of fresh collection is got in aseptic operating platform 1g after homogeneous in the enrichment medium of 50mL (250mL triangular flask) sterilizing and cultivate, culture condition is that 28 DEG C of 180rpm cultivate 72h.By bacterium liquid gradient dilution to 10 good for enrichment
-7, each dilution gradient is got 0.1mL and is spread evenly across on screening culture medium flat board, 28 DEG C of constant temperature culture.Be extracted in the dull and stereotyped upper single bacterium colony (Fig. 1) producing hydrolysis circle of screening, line picking mono-clonal in seed culture medium 28 DEG C, 180rpm shakes cultivation and is inoculated in fermentation culture in fermention medium in 12 hours.Fermented and obtained its supernatant by centrifugal (4 DEG C, the centrifugal 10min of 11000g) and detected Phospholipase D hydrolytic activity, the hydrolysis vigor of OUC-Qa2 is 0.336U/mL after measured.
Enrichment medium: 10g/L peptone, 3g/L NaCl, 0.5g/L MgSO
47H
2o, 1g/L CaCl
2, after pH 6.5,121 DEG C of steam sterilizing 20min cool, add the fresh yolk of 10mL.
Screening culture medium: 10g/L peptone, 3g/L NaCl, 0.5g/L MgSO
47H
2o, 1g/L CaCl
2, 20g/L agar, pH 6.5,121 DEG C of steam sterilizings, add the fresh yolk of 20mL after 20min cooling.
Seed culture medium: 10g/L peptone, 5g/L yeast powder, 10g/L NaCl, pH 7.4,121 DEG C of vapor sterilization 20min.
Fermention medium: 10g/L peptone, 3g/L NaCl, 0.5g/L MgSO
47H
2o, 1g/L CaCl
2, after pH 6.5,121 DEG C of steam sterilizing 20min cool, add 2% fresh yolk.
(2) qualification of OUC-Qa2
Utilize TIANGEN bacterium full genome to extract the gene of test kit extraction OUC-Qa2, by 16S rDNA, Bacteria Identification is carried out to it.Its 16S PCR primer is checked order, by sequencing result in NCBI comparison, finds that the bacterial strain the most similar to OUC-Qa2 is Acinetobacter radioresistens WC-A-157 (NCBI numbering JN669146.1).Find that OUC-Qa2 and Acinetobacter radioresistens is in evolution the most close (Fig. 2) simultaneously.
This bacterial strain was deposited in China Committee for Culture Collection of Microorganisms's common micro-organisms center (the China General Microbiological Culture Collection Center being positioned at No. 3 Institute of Microorganism, Academia Sinica in Yard 1, BeiChen xi Road, Chaoyang District, Beijing City on 01 30th, 2015, CGMCC), deposit number is CGMCC NO.10471.
Embodiment 2 OUC-Qa2 is applied to phosphatidylserine synthesis
By the fermented liquid lyophilize of OUC-Qa2, synthetic phospholipid acyl Serine in diphasic system.Reaction system is by enzyme powder 70mg (hydrolysis vigor 23U), and pH 5.6,0.2mol/L acetate buffer solution are (containing 1mol/L Serine and 0.1mol/L CaCl
2) 1mL, 20mg/mL ether dissolution 95% Ovum Gallus domesticus Flavus lecithin 1mL, 50 DEG C of water-bath oscillatory reaction 12h.Reaction terminates rear leaving standstill and treats solution layering, upper strata ether is extracted nitrogen mutually and dries up, cross 0.22 μm of organic filter membrane, PA, PC, PS in nmr quantitative detection reaction system after n-hexane dissolution constant volume.Measurement result is shown in Fig. 3.
Integrating peak areas is carried out to PA, PS, the PC in Fig. 3, then becomes the content of PA, PS, PC in reaction system with phosphorus typical curve comparing calculation.In result display reaction system, the area ratio of PA:PS:PC is 1.01:2.87:2.06.Calculate 50 DEG C, 70mg enzyme powder (hydrolysis vigor is 23U) simultaneously, react symbiosis in 12 hours and become the amount of PS to be 13.1g, far above the vigor of other PLD of bibliographical information.
Calculate by Fig. 3 data transformation efficiency and the selectivity that OUC-Qa2 produces PLD synthesis PS, wherein synthesize the transformation efficiency=PS/ (PS+PA+PC) × 100% of PS, the transformation efficiency that calculating acquisition OUC-Qa2 product PLD synthesizes PS is 47.8%.
Selectivity=the PS (PS+PA) × 100% of synthesis PS, calculating the selectivity obtaining OUC-Qa2 product PLD synthesis PS is 74%.
The PLD enzyme of OUC-Qa2 fermentative production is utilized to carry out PS synthesis, the transformation efficiency being synthesized PS by magnetic resonance detection OUC-Qa2PLD is 47.8%, the selectivity of PS synthesis is 74%, and hydrolysis produces the amount low (26%) of PA, has very high application prospect.
Claims (4)
1. an acinetobacter calcoaceticus, is characterized in that, the deposit number of described acinetobacter calcoaceticus is CGMCCNO.10471.
2. a bacterium liquid, is characterized in that, described bacterium liquid is made up of acinetobacter calcoaceticus fermentation according to claim 1.
3. the application of acinetobacter calcoaceticus according to claim 1 in synthetic phospholipid acyl Serine.
4. produce a method for phosphatidylserine, it is characterized in that, described method carrys out synthetic phospholipid acyl Serine with acinetobacter calcoaceticus catalyze phospholipid phatidylcholine according to claim 1.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510325298.8A CN104845922B (en) | 2015-06-15 | 2015-06-15 | A kind of acinetobacter calcoaceticus and the application in phosphatidylserine synthesis thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510325298.8A CN104845922B (en) | 2015-06-15 | 2015-06-15 | A kind of acinetobacter calcoaceticus and the application in phosphatidylserine synthesis thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN104845922A true CN104845922A (en) | 2015-08-19 |
| CN104845922B CN104845922B (en) | 2016-02-17 |
Family
ID=53845914
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510325298.8A Active CN104845922B (en) | 2015-06-15 | 2015-06-15 | A kind of acinetobacter calcoaceticus and the application in phosphatidylserine synthesis thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104845922B (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2020224232A1 (en) * | 2019-05-08 | 2020-11-12 | 江南大学 | Acinetobacter and use thereof in production of chiral 3-cyclohexene-1-formic acid |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2013044076A1 (en) * | 2011-09-22 | 2013-03-28 | Codexis, Inc. | Direct biocatalytic production of acrylic acid and other carboxylic acid compounds |
| CN103421719A (en) * | 2013-08-13 | 2013-12-04 | 沈阳药科大学 | Actinomycete Streptomycesbottropensis and application thereof |
| WO2015017045A1 (en) * | 2013-07-03 | 2015-02-05 | Keclon Sa | Modified bacillus cereus phospholipase c protein and method of processing vegetable oil |
| CN104388328A (en) * | 2014-08-26 | 2015-03-04 | 河北农业大学 | Novel strain for degrading 5-ring and 6-ring polycyclic aromatic hydrocarbons, and acquisition method and application thereof |
-
2015
- 2015-06-15 CN CN201510325298.8A patent/CN104845922B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2013044076A1 (en) * | 2011-09-22 | 2013-03-28 | Codexis, Inc. | Direct biocatalytic production of acrylic acid and other carboxylic acid compounds |
| WO2015017045A1 (en) * | 2013-07-03 | 2015-02-05 | Keclon Sa | Modified bacillus cereus phospholipase c protein and method of processing vegetable oil |
| CN103421719A (en) * | 2013-08-13 | 2013-12-04 | 沈阳药科大学 | Actinomycete Streptomycesbottropensis and application thereof |
| CN104388328A (en) * | 2014-08-26 | 2015-03-04 | 河北农业大学 | Novel strain for degrading 5-ring and 6-ring polycyclic aromatic hydrocarbons, and acquisition method and application thereof |
Non-Patent Citations (2)
| Title |
|---|
| VIDAR LEHMANN: "THE NATURE OF PHOSPHOLIPASE C FROM ACINETOBACTER CALCOACETICUS:EFFECTS ON WHOLE RED CELLS AND RED CELL MEMBRANES", 《ACTA PATH. MICROBIOL. SCAND.》, vol. 81, 31 December 1973 (1973-12-31), pages 419 - 426 * |
| 胡飞等: "生物酶法制备磷脂酰丝氨酸的研究进展", 《中国油脂》, vol. 37, no. 6, 31 December 2012 (2012-12-31), pages 54 - 58 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2020224232A1 (en) * | 2019-05-08 | 2020-11-12 | 江南大学 | Acinetobacter and use thereof in production of chiral 3-cyclohexene-1-formic acid |
| US11441133B2 (en) | 2019-05-08 | 2022-09-13 | Jiangnan University | Acinetobacter and use thereof in production of chiral 3-cyclohexene-1-carboxylic acid |
Also Published As
| Publication number | Publication date |
|---|---|
| CN104845922B (en) | 2016-02-17 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Tang et al. | Actinopolyspora alba sp. nov. and Actinopolyspora erythraea sp. nov., isolated from a salt field, and reclassification of Actinopolyspora iraqiensis Ruan et al. 1994 as a heterotypic synonym of Saccharomonospora halophila | |
| Xing et al. | Nocardia endophytica sp. nov., an endophytic actinomycete isolated from the oil-seed plant Jatropha curcas L. | |
| CN102925504B (en) | Method and fermentation culture medium used for synthesizing gamma-aminobutyric acid through microbial fermentation | |
| CN104087511A (en) | Mucor racemosus strain and its application | |
| Lech | Optimisation of protein-free waste whey supplementation used for the industrial microbiological production of lactic acid | |
| CN107384844A (en) | A kind of recombination bacillus coli for producing phospholipase D and its application | |
| Fan et al. | Novel millet-based flavored yogurt enriched with superoxide dismutase | |
| CN106282041A (en) | One plant height produces the bacillus subtilis mutant strain of surface element and utilizes this mutant to carry out the method that semi-solid ferment produces surface element | |
| CN107137628A (en) | Hyaluronidase inhibitor and its application from symbiotic fermentation product | |
| Wikandari et al. | Correlations between the chemical, microbiological characteristics and sensory profile of fungal fermented food | |
| Ma et al. | Intracellular response of Bacillus natto in response to different oxygen supply and its influence on menaquinone-7 biosynthesis | |
| CN101268998A (en) | Active microecological herb amino acid complex and preparation method thereof | |
| Wang et al. | Solid-state fermentation of soybean meal with edible mushroom mycelium to improve its nutritional, antioxidant capacities and physicochemical properties | |
| Skrzypczak et al. | Spontaneously fermented fruiting bodies of Agaricus bisporus as a valuable source of new isolates of lactic acid bacteria with functional potential | |
| CN104254598A (en) | Bacteria and the uses thereof | |
| Long et al. | Description of a Sulfitobacter strain and its extracellular cyclodipeptides | |
| CN104845922B (en) | A kind of acinetobacter calcoaceticus and the application in phosphatidylserine synthesis thereof | |
| Gong et al. | Enhanced DPPH radical scavenging activity and Enriched γ-Aminobutyric acid in mulberry juice fermented by the probiotic Lactobacillus Brevis S3 | |
| CN104711217B (en) | A kind of biologically active peptide of new promotion lactobacter growth | |
| CN103421719A (en) | Actinomycete Streptomycesbottropensis and application thereof | |
| CN103275901B (en) | Space-induced efficient bacillus natto, application of bacillus natto and preparation method of troche of bacillus natto | |
| Jang et al. | Antidiabetic, anticholesterol, and antioxidant activity of Gryllus bimaculatus fermented by Bacillus and Lactobacillus strains | |
| Tolpeznikaite et al. | Changes in Spirulina’s Physical and Chemical Properties during Submerged and Solid-State Lacto-Fermentation | |
| Xing et al. | Pseudonocardia nantongensis sp. nov., a novel endophytic actinomycete isolated from the coastal halophyte Tamarix chinensis Lour | |
| CN114634890B (en) | Nitrogen fixing helicobacter, and separation method and application thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| EXSB | Decision made by sipo to initiate substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant |