CN102154117A - Actinomucor elegans ZGB1 and application thereof in microbial ATP synthesis - Google Patents
Actinomucor elegans ZGB1 and application thereof in microbial ATP synthesis Download PDFInfo
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- CN102154117A CN102154117A CN 201010615701 CN201010615701A CN102154117A CN 102154117 A CN102154117 A CN 102154117A CN 201010615701 CN201010615701 CN 201010615701 CN 201010615701 A CN201010615701 A CN 201010615701A CN 102154117 A CN102154117 A CN 102154117A
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Abstract
The invention provides a new strain-Actinomucor elegans ZGB1 capable of synthesizing ATP and application thereof. The strain is preserved in China general microbiological culture Collection center, and the address is as follows: the microbial research institute of western road 1, 3, national academy of sciences, north-south, morning-yang, Beijing, zip code: 100101, accession number: CGMCC No.4313, preservation date: 11/8/2010. The invention has the following beneficial effects: the strain can be used for synthesizing ATP by taking adenine as a precursor, has the advantages of low price of raw materials, rich sources, simple raw materials of a thallus culture medium, easy separation of thallus and the like, and has high conversion efficiency: ATP can be produced at a concentration of 10g/L or more in a reaction solution containing adenine at a concentration of 3g/L or more.
Description
(1) technical field
The present invention relates to a strain and can synthesize the new bacterial strain of ATP--graceful radiation Mucor (Actinomucor elegans) ZGB1, and the application in the synthetic ATP of microorganism.
(2) background technology
ATP is an adenosine triphosphate, as a kind of biochemical drug, has been used to amyotrophy, myocardial infarction, hepatitis etc. and various first aid treatment of conditions or assisting therapy.
The main both at home and abroad at present microbe transformation method production ATP that adopts, it is two kinds of main production methods of precursor raw material that the adenosine of employing and VITAMIN B4 are specifically arranged.The method of the synthetic ATP of microbial transformation that the open employing VITAMIN B4 of reporting is a precursor raw material mainly contains: Japan consonance fermentation company adopts product ammonia bacillus (Cornebacterium ammoniagenes) to transform synthetic ATP (Biosci.Boitechnol.Biochem., 65 (3), 644～650,2001); Domestic Zhongshan University adopts cereuisiae fermentum to transform synthetic ATP (Zhongshan University's journal natural science edition, 3,15～26,1975); Zhan Guyu etc. adopt mucor aromaticus Poyah 7 strain mucormycosiss such as (Mucor armaticus) to transform synthetic ATP (Northwest University's journal natural science edition, 02,118～129,1980) also be not that precursor raw material adopts graceful radiation mucor strain to transform document or the patent report of synthetic ATP so far etc., with the VITAMIN B4.
VITAMIN B4 has adopted the chemical synthesis mass production at present, therefore adopting VITAMIN B4 is the production method of precursor raw material, have that cost of material is lower, the theoretical yield of abundant, the synthetic ATP in source is than advantages such as height, transforming synthetic ATP with Mucor, also to have a yeast culture based raw material simple, advantages such as thalline separate easily.
(3) summary of the invention
The object of the invention provides the new bacterial strain that can synthesize ATP that a strain screens--graceful radiation Mucor (Actinomucor elegans) ZGB1, and the application in the synthetic ATP of microorganism.
The technical solution used in the present invention is:
Graceful radiation Mucor (Actinomucor elegans) ZGB1, be preserved in China Committee for Culture Collection of Microorganisms common micro-organisms center, address: No. 3 Institute of Microorganism, Academia Sinica in Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, postcode: 100101, deposit number: CGMCC No.4313, preservation date: on November 8th, 2010.
The present application people shakes bottle by spawn culture and transforms, and measures the method for ATP content in the conversion fluid then with paper electrophoresis method, when screening has the microorganism of synthetic ATP ability, has screened this bacterial classification.On according to bases such as flat-plate bacterial colony feature, mycelia form, conidial fructification and spore shape to its evaluation, in conjunction with 18S rDNA sequential analysis contrast (order-checking company:, determine that this bacterium is graceful radiation Mucor (Actinomucor elegans) Sangon Biotech (Shanghai) Co., Ltd.).So far do not see that having with the VITAMIN B4 is that precursor raw material adopts graceful radiation mucor strain to transform document or the patent report of synthetic ATP.
(SEQ ID No.1) is as follows for the 18S rDNA partial sequence of bacterial strain of the present invention:
TAGGATTCCTCGTTGAGAGCAATAATTACAATGCTCTATCCCCAGCACGATGAAGTTTCAAAAGTTTACCCAGACCTTCCGGCCAAGGTTATAAACTCGCTGACTTCATCAGTGTAGCGCGCGTGCGGCCCAGAACATCTAAGGGCATCACAGACCTGTTATTGCCTAAAACTTCCTTCGGCTTGAAGCCGATAGTCTCTCTAAGAAGCCAGATAGATAAATCTATTCACCAACTAAAAAGTTGGCCTGCCTATTTAGCAGAATAAGGTCTCGTTCGTTATCGGAATTAACCAGACAAATCACTCCACGAACTAAGAACGGCCATGCACCACCACCCATAGAATCTAGAAAGAGCTTTCAATCTGTCAATCCTTACTATGTCTGGACCTGGTGAGTTTCCCCGTGTTGAGTCAAATTAAGCCGCAGGCTCCACTCCTGGTGGTGCCCTTCCGTCAATTCCTTTAAGTTTCAGCCTTGCGTCCCATACTCCCCCCAGAACCCAAAAACTTTACTTTCGCTAAGATGCTGAGCCAGTCATAATAAACAGGCCCAATCTCTAGTCGGCATAGTTTGTGGTTAAGACTACGACGGTATCTAATCGTCTTCGATCCCTTAACTTTAGACCTTGATCAATGAAAACGTCCTGGGTCAAATGCTTTCGCAGTAGTTTGTCTTCGGTCAATCCAAGAATTTCACCTCTAGCGACCAAATACAAATGCCCCCAACCGTTTCTATTCATCATTACTCAAGTTCCTGAACCAACGAAATAGGGACTAGAGTCATATTTCATTATTCCATGCTAACACATTCAAGCTTGAAAGCCTGCTTTAAACACTCTGATTTGCTCATGGTAATCATTTGGCTAGAGACTATAGGCGACCGAAGCCACCCATAGCATAACCAAATACAAAAGCCTCGGCAGAAGCGAGCTTGATCAATGAAGACCAGGCCACCTCGCCTAAAGGCTAAAGTTTGACTACGGACGTTTTAACTGCAACAACTTTAATATACGCTATTGGAGCTGGAATTACCGCGGCTGCTGGCACCAGACTTGCCCTCCAATTGATCCTCGTTAAGGGATTTAAATTGTACTCATTCCAATTGCAAGACCTTAAAGGCCCTGCATTGTTATTTATTGTCACTACCTCACCATGTCGGTATTGGGTAATTTGCGCGCCTGCTGCCTTCCTTGGATGTGGTAGCCGTTTCTCAGGCTCCCTCTCCGGAATCGAACCCTAATTCCCCGTTACCCGTTGTAGGCATTGTAAGCCTCTATCTTACAATCTCAACCATGATAGGGCAGAAAATCGAGTGGACCGTCGCCAGCACAAGGCCATGCGATCCGCTTAATTATTATGAATCACCAAGGGAAACTGGTTTTACCCAGCGTTGGCTTTATCTAATAAGTGCACCCCTTCCGAAGTCAGGGCTTTTTTGCATGTATTAGCTCTAGAATTACCACGGTTATCCAAGTAGTAAAGAATATGTCACGTAGATCATAACTGATTAATGAGCCATCGCA。
ZGB1 strain morphology feature and cultural method:
1. morphological specificity
Colonial morphology (approximate referring to Fig. 1～Fig. 4) with general Mucor, flocculence, aerial hyphae highly surpasses 1cm, the initial stage is a pure white, after transfer lark gradually to, stolon and underdeveloped rhizoid are arranged, sporangiophore is upright, diameter 10.5～26.3 μ m.There is a bigger sporocyst on the major branch top, and it often has 3～8 verticillate branches down, and all there is sporocyst on each branch top.Spherical in shape or the foreign pyriform of sporocyst, surperficial fine hair shape is chocolate after the maturation, not of uniform size, diameter 30～70 μ m.Cyst wall is easily cleared up, and the capsule neck is arranged, columella circle, oval or pyriform, and smooth surface, size is uneven, diameter (25.0～60.0) * (20.5～55.0) μ m, big person can reach 67.3～66.1 μ m.Sporangiospore is short avette and spherical, slightly transparent, smooth surface, and size is uneven, and size is (10.5～21.0) * (7.9～24.0) μ m, and big person can reach 28.9 * 28.9 μ m.Sporangiospore expands when sprouting, and is typical one pole and sprouts.Can form chlamydospore.
28 ℃ of this bacterium optimum growth temperatures, 37 ℃ of growths are suppressed, 40 ℃ of growths hardly.
2. the substratum of Cai Yonging
Solid slant culture base 1: potato, glucose agar medium (PDA).
Solid slant culture base 2: glucose 20g/L, peptone 10g/L, yeast extract paste 5g/L, NaCl 5g/L, agar 20g/L, solvent are water, pH 6.0～6.5.
Liquid seed culture medium: glucose 20g/L, peptone 10g/L, yeast extract paste 5g/L, NaCl 5g/L, solvent are water, pH 6.0～6.5.
Fermention medium: glucose 30～50g/L, yeast extract paste 1～6g/L, corn steep liquor 5～20g/L, MgSO
47H
2O 0.5～2g/L, FeSO
47H
2O 0.05～0.5g/L, NH
4Cl 1～5g/L, K
2HPO
43H
2O 1～5g/L, solvent are water, and pH 6.0～6.5.
3. culture condition
Make spore suspension through the slant strains that 28 ℃, 3～4d are cultivated, insert in the liquid seed culture medium, place 28 ℃, cultivate 12h in the rotary shaking table of 160r/min with the inoculum size of 10% (V/V).Inoculum size with 10% (V/V) inserts liquid seeds liquid in the fermention medium, places 28 ℃, cultivates 12h, fermentation ends in the rotary shaking table of 160r/min.
The invention still further relates to the application of described graceful radiation Mucor ZGB1 in the synthetic ATP of microorganism.
Specifically, described being applied as: with the VITAMIN B4 is substrate, and the thalline that obtains with grace radiation Mucor ZGB1 fermentation is the enzyme source, carries out conversion reaction 6～24 hours under 25～35 ℃, and ATP product content can reach more than the 10g/L in the conversion fluid of reaction end back.Routinely from hand over, concentrate, separations purification process processing such as crystallization can obtain described product A TP.
Preferably, described application method is as follows:
(1) graceful radiation Mucor ZGB1 is seeded to fermention medium, cultivates 12～18 hours for 28～30 ℃, and fermented liquid separated and collected thalline cleans, filters and did 40～80 ℃ of freeze-day with constant temperature in back 2～4 hours, obtains graceful radiation Mucor ZGB1 thalline; Described fermention medium final concentration consists of: glucose 30～50g/L, yeast extract paste 1～6g/L, corn steep liquor 5～20g/L, MgSO
47H
2O 0.5～2g/L, FeSO
47H
2O 0.05～0.5g/L, NH
4Cl 1～5g/L, K
2HPO
43H
2O 1～5g/L, pH 6.0～6.5; The thalline that fermented liquid after the fermentation ends is collected after filtration (is 40～80 ℃ of drying treatment 2～4h) of thermostatic drying chamber through penetratingization processing earlier.Because it is very convenient that mucormycosis filter to be collected thalline, uses this penetratingization treatment process to have characteristics easy and simple to handle, that activity of conversion is high,, also has the advantage of environmental protection owing to need in conversion reaction liquid, not add treatment process such as dimethylbenzene;
(2) get step (1) graceful radiation Mucor ZGB1 thalline and substrate preparation conversion reaction liquid, carry out conversion reaction 6～24 hours under 30～35 ℃, reaction finishes the back conversion fluid and obtains described ATP through aftertreatment; Described conversion reaction liquid initial composition is as follows: glucose 60～90g/L, Na
2HPO
412H
2O 30～90g/L, MgCl
26H
2O3～6g/L, VITAMIN B4 2～6g/L, Mucor thalline 50～100g/L.Behind the reaction 6h, follow the tracks of the conversion situation that detects, termination reaction in good time with paper electrophoresis method every 2h.ATP in the reaction solution can reach more than the 10g/L, and existing mucormycosis transforms the highest ATP productive rate of bibliographical information about 8g/L.
Conversion reaction product analysis and method of calculation:
Sampling 5～20 μ L, point sample are damping fluid with the citric acid-sodium citrate solution of pH 2.5 on the wide common filter paper of 20mm, and voltage 400V, electrophoresis time are 30～60min.After electrophoresis finishes, dry up filter paper with hair dryer, under uv analyzer, retouch out product spot with standard A TP spot same position, spot is cut into slice about 2mm, with the HCl solution of 5ml 0.01mol/L at 28 ℃, lixiviate 4h in the rotary shaking table of 160r/min measures vat liquor at the absorbancy (OD of 260nm place
260), calculate the ATP productive rate by the mole specific absorbance.
ATP calculation of yield formula:
ATP productive rate=OD
260Mr
ATPC/E
ATP
In the formula: Mr
ATPBe the ATP molecular weight, C is an extension rate, E
ATPBe the ATP molar absorptivity.
Transformation efficiency equals molar yield, and numerical value equals the ratio of actual ATP productive rate and theoretical ATP productive rate.
Beneficial effect of the present invention is mainly reflected in: screening obtains the new bacterial strain that a strain can be synthesized ATP--graceful radiation Mucor (Actinomucor elegans) ZGB1, utilize this bacterial strain VITAMIN B4 to be the synthetic ATP of precursor, have advantages such as cost of material is lower, the source is abundant, the yeast culture based raw material is simple, thalline separate easily, and the transformation efficiency height: the ATP more than the 10g/L in the reaction solution that contains the above VITAMIN B4 of 3g/L, can be produced.
(4) description of drawings
Fig. 1 is a bacterial strain vegetative hyphae electromicroscopic photograph of the present invention (10 * 4 times)
Fig. 2 bacterial strain aerial hyphae of the present invention electromicroscopic photograph (10 * 4 times)
Fig. 3 bacterial strain of the present invention is the branched sporangiophore electromicroscopic photograph of total shape (10 * 10 times)
The sporocyst electromicroscopic photograph (10 * 40 times) that Fig. 4 bacterial strain cyst wall of the present invention begins to melt.
(5) embodiment
The present invention is described further below in conjunction with specific embodiment, but protection scope of the present invention is not limited in this:
Embodiment 1:
Graceful radiation Mucor ZGB1 (CGMCC No.4313) is seeded to the PDA slant medium, cultivates 3～4d for 28 ℃, makes spore suspension (about 10 with the physiological saline washing
6～10
7Cfu/mL);
Spore suspension inserts liquid seed culture medium (glucose 20g/L, peptone 10g/L, yeast extract paste 5g/L with the inoculum size of 10% (V/V), NaCl 5g/L, solvent are water, and pH 6.0～6.5) in, place 28 ℃, cultivate 12h in the rotary shaking table of 160r/min, obtain liquid seeds liquid;
Liquid seeds liquid inserts fermention medium (glucose 40g/L, yeast extract paste 3g/L, corn steep liquor 10g/L, MgSO with the inoculum size of 10% (V/V)
47H
2O 1g/L, FeSO
47H
2O 0.2g/L, NH
4Cl 3g/L, K
2HPO
43H
2O 3g/L, solvent are water, and pH 6.0～6.5) in, place 28 ℃, cultivate 12h, fermentation ends in the rotary shaking table of 160r/min.
Fermented liquid is collected thalline through suction filtration, and does with the aseptic water washing filter, and the filter dry mycelium passes through penetratingization of the cell processing of 65 ℃ of dry 3h of thermostatic drying chamber again, and it is standby to get the Mucor thalline.
Conversion reaction liquid is formed: glucose 90g/L, Na
2HPO
412H
2O 90g/L, MgCl
26H
2O 4g/L, VITAMIN B4 3g/L, Mucor thalline 80g/L.
The conversion reaction liquid of above-mentioned preparation carries out conversion reaction in 33 ℃, the rotary shaking table of 160r/min, the output in ATP2Na during 10h is 11.14g/L, and molar yield can reach 91.07%.
Embodiment 2:
Graceful radiation Mucor ZGB1 (CGMCC No.4313) solid slant culture base (glucose 20g/L, peptone 10g/L, yeast extract paste 5g/L, NaCl 5g/L, agar 20g/L, solvent are water, pH 6.0～6.5), cultivate 3～4d for 28 ℃, make spore suspension (about 10 with the physiological saline washing
6～10
7Cfu/mL);
Spore suspension inserts liquid seed culture medium (glucose 20g/L, peptone 10g/L, yeast extract paste 5g/L with the inoculum size of 10% (V/V), NaCl 5g/L, solvent are water, and pH 6.0～6.5) in, place 28 ℃, cultivate 12h in the rotary shaking table of 160r/min, obtain liquid seeds liquid;
Liquid seeds liquid inserts fermention medium (glucose 40g/L, yeast extract paste 3g/L, corn steep liquor 10g/L, MgSO with the inoculum size of 10% (V/V)
47H
2O 1g/L, FeSO
47H
2O 0.2g/L, NH
4Cl 3g/L, K
2HPO
43H
2O 3g/L, solvent are water, and pH 6.0～6.5) in, place 28 ℃, cultivate 12h, fermentation ends in the rotary shaking table of 160r/min.
Fermented liquid is collected thalline through suction filtration, and does with the aseptic water washing filter, and the filter dry mycelium passes through penetratingization of the cell processing of 60 ℃ of dry 3h of thermostatic drying chamber again, and it is standby to get the Mucor thalline.
Conversion reaction liquid: glucose 90g/L, Na
2HPO
412H
2O 90g/L, MgCl
26H
2O 4g/L, VITAMIN B4 4g/L, Mucor thalline 80g/L.
Above-mentioned conversion reaction liquid carries out conversion reaction in 33 ℃, the rotary shaking table of 160r/min, ATP2Na output is 13.64g/L during reaction 20h, the molar yield 83.63% of ATP2Na.
SEQUENCE LISTING
＜110〉Zhejiang Polytechnical University
＜120〉graceful radiation Mucor ZGB1 and the application in the synthetic ATP of microorganism thereof
<130>
<160> 1
<170> PatentIn version 3.4
<210> 1
<211> 1522
<212> DNA
<213> Actinomucor elegans
<400> 1
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gcgtgcggcc cagaacatct aagggcatca cagacctgtt attgcctaaa acttccttcg 180
gcttgaagcc gatagtctct ctaagaagcc agatagataa atctattcac caactaaaaa 240
gttggcctgc ctatttagca gaataaggtc tcgttcgtta tcggaattaa ccagacaaat 300
cactccacga actaagaacg gccatgcacc accacccata gaatctagaa agagctttca 360
atctgtcaat ccttactatg tctggacctg gtgagtttcc ccgtgttgag tcaaattaag 420
ccgcaggctc cactcctggt ggtgcccttc cgtcaattcc tttaagtttc agccttgcgt 480
cccatactcc ccccagaacc caaaaacttt actttcgcta agatgctgag ccagtcataa 540
taaacaggcc caatctctag tcggcatagt ttgtggttaa gactacgacg gtatctaatc 600
gtcttcgatc ccttaacttt agaccttgat caatgaaaac gtcctgggtc aaatgctttc 660
gcagtagttt gtcttcggtc aatccaagaa tttcacctct agcgaccaaa tacaaatgcc 720
cccaaccgtt tctattcatc attactcaag ttcctgaacc aacgaaatag ggactagagt 780
catatttcat tattccatgc taacacattc aagcttgaaa gcctgcttta aacactctga 840
tttgctcatg gtaatcattt ggctagagac tataggcgac cgaagccacc catagcataa 900
ccaaatacaa aagcctcggc agaagcgagc ttgatcaatg aagaccaggc cacctcgcct 960
aaaggctaaa gtttgactac ggacgtttta actgcaacaa ctttaatata cgctattgga 1020
gctggaatta ccgcggctgc tggcaccaga cttgccctcc aattgatcct cgttaaggga 1080
tttaaattgt actcattcca attgcaagac cttaaaggcc ctgcattgtt atttattgtc 1140
actacctcac catgtcggta ttgggtaatt tgcgcgcctg ctgccttcct tggatgtggt 1200
agccgtttct caggctccct ctccggaatc gaaccctaat tccccgttac ccgttgtagg 1260
cattgtaagc ctctatctta caatctcaac catgataggg cagaaaatcg agtggaccgt 1320
cgccagcaca aggccatgcg atccgcttaa ttattatgaa tcaccaaggg aaactggttt 1380
tacccagcgt tggctttatc taataagtgc accccttccg aagtcagggc ttttttgcat 1440
gtattagctc tagaattacc acggttatcc aagtagtaaa gaatatgtca cgtagatcat 1500
aactgattaa tgagccatcg ca 1522
Claims (4)
1. graceful radiation Mucor (Actinomucor elegans) ZGB1, be preserved in China Committee for Culture Collection of Microorganisms common micro-organisms center, address: No. 3 Institute of Microorganism, Academia Sinica in Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, postcode: 100101, deposit number: CGMCCNo.4313, preservation date: on November 8th, 2010.
2. the application of graceful radiation Mucor ZGB1 as claimed in claim 1 in the synthetic ATP of microorganism.
3. application as claimed in claim 2, it is characterized in that described being applied as: be substrate with the VITAMIN B4, the thalline that obtains with grace radiation Mucor ZGB1 fermentation is the enzyme source, carries out conversion reaction 6～24 hours under 25～35 ℃, and conversion fluid obtains described ATP through aftertreatment.
4. application as claimed in claim 2 is characterized in that described application is as follows:
(1) graceful radiation Mucor ZGB1 is seeded to fermention medium, cultivates 12～18 hours for 28～30 ℃, and fermented liquid separated and collected thalline cleans, filters and did 40～80 ℃ of freeze-day with constant temperature in back 2～4 hours, obtains graceful radiation Mucor ZGB1 thalline; Described fermention medium final concentration consists of: glucose 30～50g/L, yeast extract paste 1～6g/L, corn steep liquor 5～20g/L, MgSO
47H
2O 0.5～2g/L, FeSO
47H
2O 0.05～0.5g/L, NH
4Cl 1～5g/L, K
2HPO
43H
2O 1～5g/L, pH 6.0～6.5;
(2) get step (1) graceful radiation Mucor ZGB1 thalline and substrate preparation conversion reaction liquid, carry out conversion reaction 6～24 hours under 30～35 ℃, reaction finishes the back conversion fluid and obtains described ATP through aftertreatment; Described conversion reaction liquid initial composition is as follows: glucose 60～90g/L, Na
2HPO
412H
2O 30～90g/L, MgCl
26H
2O3～6g/L, VITAMIN B4 2～6g/L, graceful radiation Mucor ZGB1 thalline 50～100g/L.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103993055A (en) * | 2014-04-22 | 2014-08-20 | 浙江工业大学 | Biosynthesis method of ademetionine |
| CN111454848A (en) * | 2020-04-15 | 2020-07-28 | 中国科学院微生物研究所 | Actinomucor elegans GD48, fermentation product, microbial inoculum and application thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1563398A (en) * | 2004-04-05 | 2005-01-12 | 四川省时代生物技术有限责任公司 | Application of Yazhi-fangshe Mucor in use for preparing ceramide |
| CN101487002A (en) * | 2009-03-02 | 2009-07-22 | 华南理工大学 | Alkaline protease |
-
2010
- 2010-12-31 CN CN2010106157018A patent/CN102154117B/en not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1563398A (en) * | 2004-04-05 | 2005-01-12 | 四川省时代生物技术有限责任公司 | Application of Yazhi-fangshe Mucor in use for preparing ceramide |
| CN101487002A (en) * | 2009-03-02 | 2009-07-22 | 华南理工大学 | Alkaline protease |
Non-Patent Citations (4)
| Title |
|---|
| 《中国医药工业杂志》 20031231 朱家荣等 腺嘌呤用固定化酵母生产三磷腺苷的初步研究 388-390 1-4 第34卷, 第8期 2 * |
| 《工业微生物》 20030331 朱家荣等 提高毛霉菌转化合成腺三磷产率的研究 31-33 1-4 第33卷, 第1期 2 * |
| 《微生物学报》 19771231 武功 用毛霉生产核苷酸类衍生物 I.利用毛霉酶促合成三磷酸腺苷 47-51 1-4 第17卷, 第1期 2 * |
| 《西北大学学报（自然科学版）》 19801231 谢伯泰等 用毛霉生产核苷酸类衍生物--毛霉用于以腺嘌呤为底物的ATP酶促合成 118-129 1-4 , 2 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103993055A (en) * | 2014-04-22 | 2014-08-20 | 浙江工业大学 | Biosynthesis method of ademetionine |
| CN111454848A (en) * | 2020-04-15 | 2020-07-28 | 中国科学院微生物研究所 | Actinomucor elegans GD48, fermentation product, microbial inoculum and application thereof |
| CN111454848B (en) * | 2020-04-15 | 2022-04-19 | 中国科学院微生物研究所 | Actinomucor elegans GD48, fermentation product, microbial inoculum and application thereof |
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