CN1089807C - Method for producing L-aspartic acid - Google Patents
Method for producing L-aspartic acid Download PDFInfo
- Publication number
- CN1089807C CN1089807C CN98114909A CN98114909A CN1089807C CN 1089807 C CN1089807 C CN 1089807C CN 98114909 A CN98114909 A CN 98114909A CN 98114909 A CN98114909 A CN 98114909A CN 1089807 C CN1089807 C CN 1089807C
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- CN
- China
- Prior art keywords
- aspartic acid
- atcc
- microorganism
- rhodococcus
- red coccus
- 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.)
- Expired - Fee Related
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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
- C12P13/00—Preparation of nitrogen-containing organic compounds
- C12P13/04—Alpha- or beta- amino acids
- C12P13/20—Aspartic acid; Asparagine
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- Organic Chemistry (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Wood Science & Technology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Microbiology (AREA)
- General Chemical & Material Sciences (AREA)
- Biotechnology (AREA)
- Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
Production of L-aspartic acid comprises using a Rhodococcus microorganism with aspartase activity and fumaric acid and ammonia as starting materials. The Rhodococcus used is preferably Rhodococcus sp. EA4, R. rhodochrous ATCC 17041, R. rhodochrous ATCC 17895, R. rhodochrous ATCC 19140 or R. rhodochrous ATCC 33258.
Description
The present invention relates to be used by fumaric acid and ammonia efficiently a kind of method of microorganisms producing L-aspartic acid, this microorganism is Rhod (Rhodococcus) and has aspartase activity.
The L-aspartic acid is used as medicine, foodstuff additive or analogue always.Recently the L-aspartic acid has also caused attention as the material of tensio-active agent and biodegradable inner complex.
The L-aspartic acid is mainly by being used the enzymatic process production of L-Aspartase by fumaric acid and ammonia.For example, the method (Japanese unexamined patent 56-26 196) of the bacterium of method (Japanese unexamined patent 59-113 887) of the bacterium of the method for known use intestinal bacteria (Escherichia coli) (Japanese unexamined patent 60-133883), use serratia (Serratia) and use brevibacterium sp (Brevibacterium).
The objective of the invention is further to seek those has the microorganism of aspartase activity and uses these microorganisms to produce the L-aspartic acid efficiently.
As the result of thoroughgoing and painstaking in order to achieve the above object and broad research, the inventor has found some to have the microorganism of aspartase activity and Rhod that can the High-efficient Production aspartic acid.Like this, the present invention has finished.The existence of aspartase activity is not known always in the Rhod bacterium.Up to now, less than report about aspartase activity in Rhod.
The present invention relates to the method that the action of microorganisms by having aspartase activity is produced the L-aspartic acid by fumaric acid and ammonia, it is characterized in that this microorganism is the Rhod microorganism.
As Rhod microorganism of the present invention, for example can enumerate rhodococcus EA4 (Rhodococcus sp.EA4, FERM BP-6231), prunosus red coccus (Rhodococcus rhodochrous) ATCC 17041, prunosus red coccus ATCC 17895, prunosus red coccus ATCC 19140 and prunosus red coccus ATCC33258.
In these microorganisms, the EA4 bacterial classification number is deposited in state-run life science of Japanese Science and Technology Department and human body technical institute (the National Institute of Bioscience and Human-Technology, Agency of Industrial Science and Technolog) with above-mentioned go into to hide.The bacteriology characteristic of this bacterial classification has description for 892 li at Japanese unexamined patent publication number 4-304.ATCC 17041, ATCC 17895, ATCC 19140 and ATCC 33258 bacterial classifications can easily obtain from American type culture collection (ATCC).
The microorganism that the present invention uses is cultivated containing can absorb in carbon source (for example glycerine, glucose, sucrose, lactose, fructose), nitrogenous source (for example meat extract, yeast extract paste, malt extract, corn steep liquor, polypepton (polypeptone)) and the traditional substratum for the necessary inorganic salt of each microorganism growth (for example magnesium chloride, sodium sulfate, calcium chloride, manganous sulfate, iron(ic) chloride, zinc sulfate).In above-mentioned substratum.The L-aspartic acid can be used as carbon and nitrogenous source, and fumaric acid can be used as carbon source.The add-on of these materials should be respectively 0.1～5%.
The pH value of nutrient solution is in 4～10 scope.Cultivation was under aerobic conditions carried out 1～7 day under 5～50 ℃ temperature.Cultivation is performed until aspartase activity and reaches till the maximum value.
Reaction is implemented by following mode.From as the nutrient solution of above-mentioned cultured microorganism in isolated cells or a kind of water-bearing media such as water or damping fluid, suspend by handling the material (for example stem cell, somatoblast, isolating thick or pure enzyme, immobilized cell or enzyme) that this class cell obtains.Then this suspension is contacted with a kind of substrate material.
They are that the substrate material one of this reaction can join in the reaction system with various forms for fumaric acid and ammonia one.For example, can use the mixture of ammonium fumarate or fumaric acid or its salt and a kind of inorganic ammonium salt.As fumarate, sodium fumarate or Potassium fumarate are suitable for.As inorganic ammonium salt, can use ammonium chloride, ammonium sulfate, ammonium phosphate or their mixture.
The mol ratio of fumaric acid or its salt and inorganic ammonium salt is preferably in 1: 1.5～1: 2 scope.
Temperature of reaction is 0～60 ℃, preferred 20～50 ℃.Reaction was carried out 0.1～100 hour.
During reaction.The ph value adjusts to 6～10, and preferred 8～10.
Usually, with a kind of divalent-metal ion, for example calcium or magnesium ion are with 0..1～100mM, and preferred 1～10mM joins in the reaction soln.
Below with reference to following embodiment the present invention is described in more detail.But, the present invention is not limited by this embodiment.
Embodiment 1
With rhodococcus EA4, prunosus red coccus ATCC 17041, prunosus red coccus ATCC 17895, prunosus red coccus ATCC 19410 and prunosus red coccus ATCC 33258 are inoculated into 10mlMYK substratum (0.5% polypepton, 0.3% bacterium-yeast extract paste, 0.3% bacterium-malt extract, 0.2%K respectively
2HPO
4, 0.2%KH
2PO
4) lining, and 30 ℃ of cultivations 3 days.Then, this culture solution of 0.5ml is inoculated into ASP substratum (1%L-aspartic acid, 1% polypepton, 1% bacterium-yeast extract paste, the 0.5%K of 50ml
2HPO, pH7.0) in, and cultivated 96 hours at 30 ℃.After this, their cell obtains, washs and suspend in a spot of damping fluid at last with 100mM phosphoric acid buffer (pH8.0) by centrifugation.The cell damping fluid that generates ice-cooled broken down, obtains a kind of thick enzyme solution by centrifugation with supersonic method subsequently.Measure the aspartase activity of this kind of enzyme solution.
By thick enzyme solution and 1M ammonium fumarate (pH8.8 contains the 1mM magnesium chloride) contact carrying out aspartase activity is measured, and use the L-aspartic acid 20 hours that the ODS posts generate by the HPLC quantitative assay at 30 ℃.The results are shown in Table 1.
Table 1 strain L-aspartic acid (mM) rhodococcus EA4 210 prunosus red coccus ATCC 1,704 140 prunosus red coccus ATCC 17,895 50 prunosus red coccus ATCC 19,140 170 prunosus red coccus ATCC 33,258 100
By using rhodococcus deutero-L-Aspartase, can produce the L-aspartic acid by fumaric acid and ammonia.
Claims (2)
1, the method for producing the L-aspartic acid by fumaric acid and ammonia by the action of microorganisms with aspartase activity is characterized in that this microorganism is a kind of microorganism of Rhod.
2, the process of claim 1 wherein that the microorganism of Rhod is rhodococcus EA4, prunosus red coccus ATCC 17041, prunosus red coccus ATCC 17895, prunosus red coccus ATCC 19140 or prunosus red coccus ATCC 33258.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP14706097A JPH10313888A (en) | 1997-05-22 | 1997-05-22 | Production of l-aspartic acid |
| JP147060/97 | 1997-05-22 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1200405A CN1200405A (en) | 1998-12-02 |
| CN1089807C true CN1089807C (en) | 2002-08-28 |
Family
ID=15421592
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN98114909A Expired - Fee Related CN1089807C (en) | 1997-05-22 | 1998-05-22 | Method for producing L-aspartic acid |
Country Status (3)
| Country | Link |
|---|---|
| JP (1) | JPH10313888A (en) |
| CN (1) | CN1089807C (en) |
| FR (1) | FR2763599B1 (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH10337185A (en) * | 1997-06-06 | 1998-12-22 | Mitsubishi Rayon Co Ltd | New protein having aspartase activity and genetic dna encoding the same |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1125769A (en) * | 1994-04-20 | 1996-07-03 | 罗纳·布朗克化学公司 | Method for preparation L-aspartate from aspartat amonia |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US3391059A (en) * | 1964-02-19 | 1968-07-02 | Sanraku Ocean Kabushiki Kaisha | Process for producing l-aspartic acid |
| JPS5626196A (en) * | 1979-08-10 | 1981-03-13 | Mitsubishi Petrochem Co Ltd | Preparation of l-aspartic acid |
| JPH10337185A (en) * | 1997-06-06 | 1998-12-22 | Mitsubishi Rayon Co Ltd | New protein having aspartase activity and genetic dna encoding the same |
-
1997
- 1997-05-22 JP JP14706097A patent/JPH10313888A/en active Pending
-
1998
- 1998-05-22 FR FR9806455A patent/FR2763599B1/en not_active Expired - Fee Related
- 1998-05-22 CN CN98114909A patent/CN1089807C/en not_active Expired - Fee Related
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1125769A (en) * | 1994-04-20 | 1996-07-03 | 罗纳·布朗克化学公司 | Method for preparation L-aspartate from aspartat amonia |
Also Published As
| Publication number | Publication date |
|---|---|
| FR2763599A1 (en) | 1998-11-27 |
| CN1200405A (en) | 1998-12-02 |
| FR2763599B1 (en) | 2004-11-26 |
| JPH10313888A (en) | 1998-12-02 |
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Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C53 | Correction of patent for invention or patent application | ||
| CB02 | Change of applicant information |
Applicant after: Mitsubishi Rayon Co., Ltd. Applicant before: Nippon Chemical Industry Co., Ltd. |
|
| COR | Change of bibliographic data |
Free format text: CORRECT: APPLICANT; FROM: NIPPON CHEMICAL INDUSTRY CO., LTD. TO: MITSUBISHI RAYON CO., LTD. |
|
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| C17 | Cessation of patent right | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20020828 Termination date: 20100522 |