CN1005155B - Production of l-threonine by the fermentation method - Google Patents
Production of l-threonine by the fermentation method Download PDFInfo
- Publication number
- CN1005155B CN1005155B CN85104604.5A CN85104604A CN1005155B CN 1005155 B CN1005155 B CN 1005155B CN 85104604 A CN85104604 A CN 85104604A CN 1005155 B CN1005155 B CN 1005155B
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- China
- Prior art keywords
- threonine
- aec
- ahv
- brevibacterium lactofermentum
- present
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Abstract
The present invention relates to a method for producing L-threonine by a fermentation method. The present invention uses a double resistant variation strain of brevibacterium bacteria, which has alpha-amino-beta-hydroxy valeric acid (AHV) and S-(2-aminoethyl)-L-cysteine (AEC); the present invention uses the double resistance of the AHV and the AEC of brevibacterium lactofermentum, the variation strain obtained by the selection of a culture medium (a succinic acid culture medium for short) which uses succinic acid as a unique carbon source is used as L-threonine production bacteria, and the yield of the L-threonine of a representative strain C 2-1261 is 16 mg/ml. A research result indicates that the succinic acid culture medium can be effectively used for the sieving of the L-threonine production bacteria.
Description
The present invention relates to the fermentation method for producing of L-Threonine.
Producing l-threonine by fermentation, existing report adopts pantonine-hydroxypentanoic acid (AHV) and the S-(2-amino-ethyl of brevibacterium lactofermentum (Brevibacterium Lactofermentum))-the dual resistant mutant of L-halfcystine (AEC) is as L-Threonine production bacterium.The L-Threonine that the present invention uses is produced bacterium, then is from the brevibacterium lactofermentum inductive, has AHV and the dual resistance of AEC and by being that the plate isolation substratum of sole carbon source is selected (hereinafter to be referred as SAM with the succsinic acid
9, mean growing on succinicacid medium) this bacterial strain of variant brevibacterium lactofermentum C2-1261(be preserved in China Committee for Culture Collection of Microorganisms common micro-organisms center on September 25th, 1985 with the preserving number of CGMCC NO.0084).At first induce AHV resistance and AEC resistance, revest SAM from wild type strain brevibacterium lactofermentum ATCC13869
9Mark can obtain variant of the present invention.
The concrete grammar of seed selection variant of the present invention below is described.At first from ATCC13869, according to document (in gloomy luxuriant, the vertebra tail is brave: * ferment と metabolism, 32:56,1975.) described method seed selection AHV resistance and AEC resistant mutant, therefrom screening obtains brevibacterium lactofermentum B7-8354(AHV
*AEC
*).It is 10 that the thalline that is incubated at the B7-8354 of slant medium overnight is made concentration with the 0.1MpH7.2 phosphate buffered saline buffer
8The single-cell suspension liquid of individual/milliliter adds ethylmethane sulfonate (ultimate density is 0.4M), 31 ℃ of oscillation treatment 1 hour, after the stopped reaction treatment solution is coated on the succsinic acid substratum (SAM), cultivated 2～6 days for 31 ℃, select the macrocolony faster of on this substratum, growing, be SAM
9Variant.In order to test the L-Threonine throughput of each variant, with the slant culture thalline of each variant connect one encircle in the test tube that fills the 5ml fermention medium (2 * 20cm) or the 500ml triangular flask of 20ml fermention medium in, 31 ℃ of shaking culture are measured the L-threonine content in the fermentation culture after 72 hours.Fig. 1 represents to induce from B7-8354 175 strain SAM of acquisition
9The pipe that shakes of variant produces the acid distribution.Fig. 2 represents the highest strain C2-1261(AHV
*AEC
*SAM
9) and its parental plant B7-8354(AHV
*AEC
*) comparison of acid producing ability.The result shows, owing to induce SAM
9Variation increases substantially L-Threonine output.
The separated and collected of L-Threonine can adopt common ion exchange extraction technology to carry out in the fermentation culture.
The inclined-plane thalline of cultivating one day brevibacterium lactofermentum C2-1261 is connect one encircle in the 500ml triangular flask that fills the 20ml fermention medium, 31 ℃ of shaking culture 72 hours, the L-threonine content can reach 16mg/ml in the nutrient solution.Merge each shake-flask culture liquid altogether 1 liter, centrifugal removal thalline and lime carbonate, supernatant liquor flows into 732 cationic exchange resin adsorption, the weak ammonia wash-out, elutriant is concentrated through decolouring, crystallisation by cooling, obtains L-Threonine coarse crystallization 11 grams.Wipe to do through recrystallization, the smart crystallization 8 of L-Threonine that can get purity and be 98.5% or more restrains again.
Slant medium is formed (%)
Glucose 0.5 NaCl 0.5
Extractum carnis 1.0 peptones 1.0
Agar 2.0 pH 7.0
The succsinic acid substratum is formed (%) fermention medium and is formed (%)
Sodium succinate 5.0 glucose 10.0
（NH
4）
2SO
40.15 （NH
4）
2SO
43.0
KH
2PO
40.1 KH
2PO
40.15
K
2HPO
40.3 MgSO
4·7H
2O 0.04
MgSO
4·7H
2O 0.01 MnSO
4·4H
2O 0.001
MnSO
4·4H
2O 0.001 FeSO
4·7H
2O 0.001
FeSO
47H
2O 0.001 d-vitamin H (g) 20
D-vitamin H (g) 3 thiamine salt hydrochlorates (g) 30
Thiamine hydrochloride (g) 10 CaCO
33.0
Urea 0.15 pH 7.0
Agar powder 2.0
pH 7.0
Claims (2)
1, a kind of method by production of L-threonine by fermentation, it is the fermention medium cultivation L-Threonine production bacterium of main carbon, nitrogenous source that this method adopts with glucose, ammonium sulfate, by common ion exchange extraction technology separated and collected L-Threonine from fermentation culture, it is characterized in that it is brevibacterium lactofermentum (Brevibacterium Lactofermentum) C2-1261(CGMCC NO.0084 that the used L-Threonine of this method is produced bacterium then).
2, brevibacterium lactofermentum C2-1261 as claimed in claim 1 is characterized by and has pantonine-hydroxypentanoic acid resistance (AHV
*), the S-(2-amino-ethyl)-L-halfcystine resistance (AEC
*) and by with the succsinic acid being the plate isolation substratum selection (SAM of sole carbon source
9) hereditary property.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN85104604.5A CN1005155B (en) | 1985-09-25 | 1985-09-25 | Production of l-threonine by the fermentation method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN85104604.5A CN1005155B (en) | 1985-09-25 | 1985-09-25 | Production of l-threonine by the fermentation method |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN85104604A CN85104604A (en) | 1987-04-08 |
| CN1005155B true CN1005155B (en) | 1989-09-13 |
Family
ID=4793949
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN85104604.5A Expired CN1005155B (en) | 1985-09-25 | 1985-09-25 | Production of l-threonine by the fermentation method |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1005155B (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR100505797B1 (en) * | 2002-10-11 | 2005-08-04 | 씨제이 주식회사 | A Mutated Microorganism Having A Knockout fadR Gene On Chromosome And A Process For Producing L-Threonine Using Said Mutant |
| KR100608085B1 (en) * | 2004-02-05 | 2006-08-02 | 씨제이 주식회사 | A microorganism producing L-threonine having an inactivated tyrR gene, method for producing the same and method for producing L-threonine using the microorganism |
-
1985
- 1985-09-25 CN CN85104604.5A patent/CN1005155B/en not_active Expired
Also Published As
| Publication number | Publication date |
|---|---|
| CN85104604A (en) | 1987-04-08 |
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