JP3651036B2 - Method for producing nucleoside-5'-phosphate ester - Google Patents
Method for producing nucleoside-5'-phosphate ester Download PDFInfo
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- JP3651036B2 JP3651036B2 JP31599694A JP31599694A JP3651036B2 JP 3651036 B2 JP3651036 B2 JP 3651036B2 JP 31599694 A JP31599694 A JP 31599694A JP 31599694 A JP31599694 A JP 31599694A JP 3651036 B2 JP3651036 B2 JP 3651036B2
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Description
【0001】
【産業上の利用分野】
この発明はヌクレオシド−5’−燐酸エステルの製造法に関する。ヌクレオシド−5’−燐酸エステルは、調味料、医薬ならびにそれらの原料として有用である。
【0002】
【従来の技術】
ヌクレオシド−5’−燐酸エステルの生化学的な製造法としては、燐酸供与体としてパラニトロフェニル燐酸を用い、ヌクレオシドを燐酸化する方法(特公昭39−29858号)、無機燐酸を用いる方法(特公昭42−1186号)、アセチル燐酸を用いる方法(特開昭56−82098号)、アデノシン三燐酸(ATP)を使用する方法(特開昭63−230094号)が知られている。
また、本発明者らは、ポリ燐酸を用いる方法を開発している(特開昭53−56390号)。
【0003】
【発明が解決しようとする課題】
しかしながら、これらの方法にあっては使用する基質が高価であったり、不安定であったりして、安定したヌクレオシド−5’−燐酸エステルの生産を行うためには、不利であった。 また、本発明者らが開発してきたポリ燐酸を用いる方法も、その後の詳細な検討の結果、生成物としてヌクレオシド−5’−燐酸エステルのみならず、2’−、3’−ヌクレオチド異性体を副生することが判明し、実用に供するには困難であることが明らかになった。
【0004】
【課題を解決するための手段】
そこで本発明者は、安定したヌクレオシド−5’−燐酸エステルの生産を行うために種々の検討を加えた結果、ヌクレオシドならびにポリ燐酸(塩)、フェニル燐酸(塩)およびカルバミル燐酸(塩)よりなる群より選択された燐酸供与体に、弱酸性条件下(pH4.0〜6.5)にて特定の微生物を作用させることにより、2’−、3’−ヌクレオチドの副生を伴うことなく、効率的にヌクレオシド−5’−燐酸エステルを生産可能なることを発見し、本発明を完成するに至った。
【0005】
すなわち、本発明は、エンテロバクター属、エシェリヒア属、モルガネラ属、クレブシエラ属、セラチア属、クロモバクテリウム属又はセデシア属に属し、ヌクレオシドならびにポリ燐酸(塩)、フェニル燐酸(塩)およびカルバミル燐酸(塩)よりなる群より選択された燐酸供与体よりヌクレオシド−5’−燐酸エステルを生成する能力を有する微生物を、pH4.0〜6.5の条件下でヌクレオシドならびにポリ燐酸(塩)、フェニル燐酸(塩)およびカルバミル燐酸(塩)よりなる群より選択された燐酸供与体に作用させて、ヌクレオシド−5’−燐酸エステルを生成せしめ、これを採取することを特徴とするヌクレオシド−5’−燐酸エステルの製造法を提供するものである。
【0006】
【作用】
本発明において使用される微生物は、エンテロバクター属、エシェリヒア属、モルガネラ属、クレブシエラ属、セラチア属、クロモバクテリウム属またはセデシア属に属し、ヌクレオシドならびにポリ燐酸(塩)、フェニル燐酸(塩)およびカルバミル燐酸(塩)よりなる群より選択された燐酸供与体からヌクレオシド−5’−燐酸エステルを生成する能力を有する微生物であり、例えば、次のような微生物が使用される。
【0007】
Escherichia blattae JCM 1650
Serratia ficaria ATCC 33105
Klebsiella planticola IFO 14939 (ATCC 33531)
Klebsiella pneumoniae IFO 3318 (ATCC 8724)
Klebsiella terrigena IFO 14941 (ATCC 33257)
Morganella morganii IFO 3168
Enterobacter aerogenes IFO 12010
Enterobacter aerogenes IFO 13534 (ATCC 13048)
Chromobacterium fluviatile IAM 13652
Chromobacterium violaceum IFO 12614
Cedecea lapagei JCM 1684
Cedecea davisiae JCM 1685
Cedecea neteri JCM 5909
【0008】
上記の微生物は、炭素源、窒素源、無機イオン等を含む通常の栄養培地中に培養される。炭素源としては、グルコース、フラクトース、シュクロース、マルトースなどの糖類の他、クエン酸、フマール酸、グルコン酸などの有機酸、エタノール、グリセリンなどのアルコール類が使用される。窒素源としては、塩化アンモニウム、硫酸アンモニウム、硝酸アンモニウムなどのアンモニウム塩、アンモニア水、アンモニアガスが好適である。無機イオンとしては、例えば第一鉄イオン、マグネシウムイオン、マンガンイオン、燐酸イオン、カリウムイオンが、必要に応じて、添加される。さらに、必要に応じ、アミノ酸、ビタミン等またはこれらを含有する酵母エキス、ペプトン、肉エキス、コーンスティープリカー、大豆蛋白加水分解物等の有機栄養源が添加される。
【0009】
上記の微生物の培養は、常法によって行われる。例えば、培地のpHを5〜8とし、微生物を接種後、20〜40℃にて5〜72時間、好気的に培養すればよい。
【0010】
上記の微生物を、ヌクレオシドならびにポリ燐酸(塩)、フェニル燐酸(塩)およびカルバミル燐酸(塩)よりなる群より選択された燐酸供与体に作用せしめヌクレオシド−5’−燐酸エステルを得る方法としては、かくして得られた培養液、培養液から分離した微生物菌体、その乾燥菌体、菌体破砕物或いはその自己消化物などの菌体処理物を、水性媒体中にて、ヌクレオシドとポリ燐酸(塩)、フェニル燐酸(塩)およびカルバミル燐酸(塩)よりなる群より選択された燐酸供与体とを接触させればよい。この際、反応のpHを4.0〜6.5の範囲の弱酸性に調整することが必要である。
【0011】
燐酸供与体として用いられるポリ燐酸(塩)としては、ピロ燐酸、トリポリ燐酸、トリメタ燐酸、テトラメタ燐酸、ヘキサメタ燐酸、又はそれらの混合物もしくはそれらのナトリウム塩、カリウム塩、あるいはそれらの塩混合物などが、フェニル燐酸(塩)としては、フェニル燐酸ジナトリウム、フェニル燐酸ジカリウム、O,O−ジフェニル燐酸無水物、又はそれらの混合物などが、カルバミル燐酸(塩)としては、カルバミル燐酸ジナトリウム、カルバミル燐酸ジカリウム、カルバミル燐酸ジアンモニウム、カルバミル燐酸ジリチウム、又はそれらの混合物などが使用可能である。燐酸供与体の使用濃度は、燐酸受容体であるヌクレオシドの濃度によって決定される。通常、ヌクレオシドの1〜5倍量の添加が望ましい。
【0012】
使用するヌクレオシドとしては、プリンヌクレオシド類として、プリンリボシド、イノシン、グアノシン、アデノシン、キサントシン等、ピリミジンヌクレオシド類として、ウリジン、シトシン等が挙げられる。使用するヌクレオシドに対応して、5’−プリンリボチド、5’−イノシン酸、5’−グアニル酸、5’−アデニル酸、5’−キサンチル酸、5’−ウリジル酸、5’−シチジル酸などが生成する。
【0013】
反応液に添加するヌクレオシドの濃度としては、0.1〜10g/dLが望ましいが、水に難溶性のヌクレオシドを使用する場合には、硼酸あるいはジメチルスルフォオキシドなどの溶解助剤を添加するとよい。
【0014】
反応は、通常、温度20〜60℃、望ましくは30〜40℃で、pH4.0〜6.5、望ましくはpH4.5〜6.0の弱酸性側が好結果を与える。この適当なpH範囲は、従来、報告されているヌクレオシド−5’−燐酸エステルの生産方法におけるpHとは異なる範囲であって、この点にも本発明の特徴がある。
【0015】
反応には、靜置あるいは攪拌の何れの方法をも採用し得る。反応時間は、使用する微生物の活性、基質の濃度などの条件によって異なるが、1〜100時間が望ましい。
【0016】
反応液中に生成したヌクレオシド−5’−燐酸エステルの分離は、通常のイオン交換樹脂を用いる方法や、その他の通常に行われる分離方法が用いられる。
【0017】
【実施例】
以下、本発明の実施例を詳細に説明する。なお、本実施例において、原料のヌクレオシドおよび生成したヌクレオシド−5’−燐酸エステルは、高速液体カラムクロマトグラフ法(HPLC、High Performance Liquid Chromatography) により、下記の機器および条件下に分析した。
カラム:コスモジル5C18−AR(4.6×150mm)[ナカライテスク社製品]
移動層:5 mM Potassium phosphate (pH 2.8): Methanol = 95:5
流速:1.0 mL/分
温度:室温
検出:UV 245nm
【0018】
(実施例1)
ペプトン1g/dL、酵母エキス0.5 g/dL、食塩1g/dLを含有する栄養培地(pH7.0)を500mL容坂口フラスコに50mL入れ、120℃にて20分加熱殺菌した。これに、斜面培養した表1に示す微生物を接種し、30℃にて16時間培養した。かくして得られた培養液より遠心分離によって菌体を回収し、菌体を酢酸バッファー液(pH5.0)に懸濁し、菌体懸濁液を調製した。
【0019】
イノシン2g/dL、ピロ燐酸10g/dL、MgSO4 2mMを、酢酸バッファー液に溶解し、これに上記の菌体懸濁液を菌体湿重量で5g/dLとなるように添加し、pHを5.0に調整、維持しながら、30℃で5時間反応させた。生成した5’−イノシン酸量を表1に示した。 なお、何れの菌株についても、2’−イノシン酸、3’−イノシン酸の副生は、ほとんど認められなかった。
【0020】
【表1】
【0021】
(実施例2)
実施例1で調製した菌株を用い、反応液中のイノシンに代えて、グアノシンを使用して同様に反応を行った。生成した5’−グアニル酸量を表2に示した。
【0022】
【表2】
【0023】
(実施例3)
実施例1で調製した菌株懸濁液を用い、反応液中のピロ燐酸に代えて、トリポリ燐酸(試薬)または「ポリゴンP」[商品名、ポリ燐酸、千代田化学(株)製品]を用い、イノシンと反応させた。生成した5’−イノシン酸量を表3に示した。
【0024】
【表3】
【0025】
(実施例4)
実施例1で調製した菌体懸濁液を用い、pH3〜8のバッファー液中で、イノシン1g/dL、ピロ燐酸ナトリウム5g/dL、硫酸マグネシウム2mMを含む反応液を調製し、この反応液中、30℃で1時間反応した。図1に示す如く、反応に使用したEscherichia blattae JCM 1650、Serratia ficaria ATCC 33105、Klebsiella pneumoniae IFO 3318、Morganella morganii IFO 3168、Enterobacter aerogenes IFO 12010、Chromobacterium fluviatile IAM 13652、Cedecea lapagei JCM 1684の各菌株の何れもが、pH5.0〜5.5の範囲において、最も効率的に5’−イノシン酸を生成した。
【0026】
(実施例5)
実施例1にて調製した菌体懸濁液を用い、pH5.0の酢酸バッファー液中でイノシン1g/dL、ピロ燐酸ナトリウム5g/dL、硫酸マグネシウム2mMを含む反応液を調製し、この反応液中、15〜60℃において反応を行った。図2に示す如く、反応に使用したEscherichia blattae JCM 1650、Klebsiella pneumoniae IFO 3318、Morganella morganii IFO 3168、 Enterobacter aerogenes IFO 12010 、Chromobacterium fluviatile IAM 13652、 Cedecea lapagei JCM 1684の各菌株の何れもが30〜40℃の範囲で、Serratia ficaria ATCC 33105 はは55℃付近において、最も効率的に5' −イノシン酸を生成した。
【0027】
(実施例6)
実施例1で調製した菌株を用い、反応液中のイノシンに代えて、ウリジンまたはシチジンを、燐酸供与体としてフェニル燐酸ジナトリウムを使用して同様に反応を行った。生成した5’−ウリジル酸量または5’−シチジル酸量を表4に示した。
【0028】
【表4】
【0029】
(実施例7)
実施例1で調製した菌株を用い、反応液中のイノシンに代えて、ウリジンまたはシチジンを、燐酸供与体としてカルバミル燐酸ジナトリウムを使用して同様に反応を行った。生成した5’−ウリジル酸量または5’−シチジル酸量を表5に示した。
【0030】
【表5】
【0031】
【発明の効果】
ヌクレオシドならびにポリ燐酸(塩)、フェニル燐酸(塩)およびカルバミル燐酸(塩)よりなる群より選択された燐酸供与体に、ヌクレオシド−5’−燐酸エステルを生成する能力を有する微生物を作用せしめ、ヌクレオシド−5’−燐酸エステルを取得する本発明の方法は、異性体の副生を伴うことなく、安定、かつ、効率よくヌクレオシド−5’−燐酸エステルを生成し、採取し得るという効果を有する。
【図面の簡単な説明】
【図1】反応pHと5’−イノシン酸の生成量との関係を示す線図である。
【図2】反応温度と5’−イノシン酸の生成量との関係を示す線図である。[0001]
[Industrial application fields]
The present invention relates to a process for producing a nucleoside-5′-phosphate ester. Nucleoside-5′-phosphate esters are useful as seasonings, pharmaceuticals, and raw materials thereof.
[0002]
[Prior art]
As a biochemical production method of nucleoside-5′-phosphate ester, paranitrophenyl phosphoric acid is used as a phosphoric acid donor, nucleoside is phosphorylated (Japanese Patent Publication No. 39-29858), and inorganic phosphoric acid is used (specialty). No. 42-1186), a method using acetyl phosphate (Japanese Patent Laid-Open No. 56-82098), and a method using adenosine triphosphate (ATP) (Japanese Patent No. 63-230094) are known.
The present inventors have also developed a method using polyphosphoric acid (Japanese Patent Laid-Open No. 53-56390).
[0003]
[Problems to be solved by the invention]
However, these methods are disadvantageous for producing a stable nucleoside-5′-phosphate ester because the substrate used is expensive or unstable. In addition, the method using polyphosphoric acid, which has been developed by the present inventors, has been developed as a result of subsequent detailed studies, and as a product, not only nucleoside-5′-phosphate esters but also 2′-, 3′-nucleotide isomers are used. It turned out to be a by-product, and it became clear that it was difficult to put it to practical use.
[0004]
[Means for Solving the Problems]
Therefore, the present inventor has made various studies in order to produce a stable nucleoside-5′-phosphate ester. By causing a specific microorganism to act on a phosphate donor selected from the group under mildly acidic conditions (pH 4.0 to 6.5), without the by-product of 2′-, 3′-nucleotides, It was discovered that nucleoside-5′-phosphate ester can be efficiently produced, and the present invention has been completed.
[0005]
That is, the present invention belongs to the genus Enterobacter, Escherichia, Morganella, Klebsiella, Serratia, Chromobacterium, or Cedesia, and includes nucleosides and polyphosphates (salts), phenylphosphates (salts), and carbamyl phosphates (salts). ) A microorganism having the ability to produce a nucleoside-5′-phosphate ester from a phosphate donor selected from the group consisting of nucleosides and polyphosphates (salts), phenylphosphates (pH) of pH 4.0-6.5. Salt) and carbamyl phosphate (salt) to act on a phosphate donor selected from the group consisting of nucleoside-5′-phosphate ester and collecting it. The manufacturing method of this is provided.
[0006]
[Action]
Microorganisms used in the present invention belong to the genus Enterobacter, Escherichia, Morganella, Klebsiella, Serratia, Chromobacterium or Cedesia, and include nucleosides and polyphosphates (salts), phenylphosphates (salts) and carbamyls. A microorganism having the ability to produce a nucleoside-5′-phosphate ester from a phosphate donor selected from the group consisting of phosphoric acid (salts). For example, the following microorganisms are used.
[0007]
Escherichia blattae JCM 1650
Serratia ficaria ATCC 33105
Klebsiella planticola IFO 14939 (ATCC 33531)
Klebsiella pneumoniae IFO 3318 (ATCC 8724)
Klebsiella terrigena IFO 14941 (ATCC 33257)
Morganella morganii IFO 3168
Enterobacter aerogenes IFO 12010
Enterobacter aerogenes IFO 13534 (ATCC 13048)
Chromobacterium fluviatile IAM 13652
Chromobacterium violaceum IFO 12614
Cedecea lapagei JCM 1684
Cedecea davisiae JCM 1685
Cedecea neteri JCM 5909
[0008]
The microorganism is cultured in a normal nutrient medium containing a carbon source, a nitrogen source, inorganic ions, and the like. As the carbon source, sugars such as glucose, fructose, sucrose and maltose, organic acids such as citric acid, fumaric acid and gluconic acid, and alcohols such as ethanol and glycerin are used. As the nitrogen source, ammonium salts such as ammonium chloride, ammonium sulfate, and ammonium nitrate, aqueous ammonia, and ammonia gas are suitable. As inorganic ions, for example, ferrous ions, magnesium ions, manganese ions, phosphate ions, and potassium ions are added as necessary. Furthermore, organic nutrient sources such as yeast extract, peptone, meat extract, corn steep liquor and soy protein hydrolyzate containing amino acids, vitamins, etc. or these are added as necessary.
[0009]
The culture of the above microorganisms is performed by a conventional method. For example, the pH of the medium may be 5 to 8, and after inoculation with microorganisms, the medium may be cultured aerobically at 20 to 40 ° C. for 5 to 72 hours.
[0010]
As a method for obtaining the nucleoside-5′-phosphate ester by causing the above-mentioned microorganism to act on a nucleoside and a phosphate donor selected from the group consisting of polyphosphoric acid (salt), phenylphosphoric acid (salt) and carbamylphosphoric acid (salt). In the aqueous medium, nucleoside and polyphosphoric acid (salts) are obtained in an aqueous medium, such as the culture broth thus obtained, microbial cells separated from the culture broth, dried microbial cells, crushed microbial cells or self-digested microbial cells. ), A phosphoric acid donor selected from the group consisting of phenylphosphoric acid (salt) and carbamylphosphoric acid (salt). At this time, it is necessary to adjust the pH of the reaction to weak acidity in the range of 4.0 to 6.5.
[0011]
Examples of the polyphosphoric acid (salt) used as the phosphoric acid donor include pyrophosphoric acid, tripolyphosphoric acid, trimetaphosphoric acid, tetrametaphosphoric acid, hexametaphosphoric acid, or a mixture thereof, or a sodium salt, potassium salt, or a salt mixture thereof. Examples of phenyl phosphoric acid (salt) include disodium phenyl phosphate, dipotassium phenyl phosphate, O, O-diphenyl phosphoric anhydride, or a mixture thereof. Examples of carbamyl phosphoric acid (salt) include disodium carbamyl phosphate, dipotassium carbamyl phosphate, Diammonium carbamyl phosphate, dilithium carbamyl phosphate, or a mixture thereof can be used. The concentration of the phosphate donor used is determined by the concentration of the nucleoside that is the phosphate acceptor. Usually, addition of 1 to 5 times the amount of nucleoside is desirable.
[0012]
Examples of the nucleoside used include purine nucleosides such as purine riboside, inosine, guanosine, adenosine and xanthosine, and pyrimidine nucleosides such as uridine and cytosine. Corresponding to the nucleoside used, 5′-purine ribotide, 5′-inosinic acid, 5′-guanylic acid, 5′-adenylic acid, 5′-xanthylic acid, 5′-uridylic acid, 5′-cytidylic acid, etc. Generate.
[0013]
The concentration of the nucleoside added to the reaction solution is preferably 0.1 to 10 g / dL. However, when using a nucleoside that is hardly soluble in water, a solubilizing agent such as boric acid or dimethyl sulfoxide may be added. .
[0014]
The reaction is usually performed at a temperature of 20 to 60 ° C., preferably 30 to 40 ° C., and a slightly acidic side having a pH of 4.0 to 6.5, preferably pH 4.5 to 6.0 gives good results. This appropriate pH range is different from the pH in the conventionally reported methods for producing nucleoside-5′-phosphate esters, and this point is also a feature of the present invention.
[0015]
For the reaction, any method of placing or stirring can be employed. The reaction time varies depending on conditions such as the activity of the microorganism used and the concentration of the substrate, but is preferably 1 to 100 hours.
[0016]
For separation of the nucleoside-5′-phosphate ester produced in the reaction solution, a method using a normal ion exchange resin or other conventional separation methods is used.
[0017]
【Example】
Hereinafter, embodiments of the present invention will be described in detail. In this Example, the raw material nucleoside and the produced nucleoside-5′-phosphate ester were analyzed by the high performance liquid chromatography (HPLC) under the following equipment and conditions.
Column: Cosmosil 5C 18 -AR (4.6 x 150 mm) [Nacalai Tesque product]
Moving layer: 5 mM Potassium phosphate (pH 2.8): Methanol = 95: 5
Flow rate: 1.0 mL / min Temperature: Room temperature Detection: UV 245 nm
[0018]
(Example 1)
50 mL of nutrient medium (pH 7.0) containing 1 g / dL of peptone, 0.5 g / dL of yeast extract, and 1 g / dL of sodium chloride was placed in a 500 mL Sakaguchi flask and sterilized by heating at 120 ° C. for 20 minutes. This was inoculated with the microorganisms shown in Table 1 on the slope culture and cultured at 30 ° C. for 16 hours. The cells were collected from the culture solution thus obtained by centrifugation, and the cells were suspended in an acetate buffer solution (pH 5.0) to prepare a cell suspension.
[0019]
Inosine 2 g / dL, pyrophosphoric acid 10 g / dL, MgSO 4 2 mM are dissolved in an acetic acid buffer solution, and the above cell suspension is added to this so that the wet cell weight becomes 5 g / dL, and the pH is adjusted. The mixture was reacted at 30 ° C. for 5 hours while adjusting and maintaining at 5.0. The amount of 5′-inosinic acid produced is shown in Table 1. In any of the strains, almost no 2′-inosinic acid or 3′-inosinic acid by-product was observed.
[0020]
[Table 1]
[0021]
(Example 2)
Using the strain prepared in Example 1, the reaction was carried out in the same manner using guanosine instead of inosine in the reaction solution. The amount of 5′-guanylic acid produced is shown in Table 2.
[0022]
[Table 2]
[0023]
(Example 3)
Using the strain suspension prepared in Example 1, instead of pyrophosphoric acid in the reaction solution, using tripolyphosphoric acid (reagent) or “polygon P” [trade name, polyphosphoric acid, product of Chiyoda Chemical Co., Ltd.] Reacted with inosine. The amount of 5′-inosinic acid produced is shown in Table 3.
[0024]
[Table 3]
[0025]
(Example 4)
Using the bacterial cell suspension prepared in Example 1, a reaction solution containing 1 g / dL inosine, 5 g / dL sodium pyrophosphate, and 2 mM magnesium sulfate in a buffer solution having a pH of 3 to 8 was prepared. , Reacted at 30 ° C. for 1 hour. As shown in FIG. 1, Escherichia blattae JCM 1650, Serratia ficaria ATCC 33105, Klebsiella pneumoniae IFO 3318, Morganella morganii IFO 3168, Enterobacter aerogenes IFO 12010, Chromobacterium fluviatile IAM 13652, and Cedecea lapagei JCM 1 Produced 5′-inosinic acid most efficiently in the range of pH 5.0 to 5.5.
[0026]
(Example 5)
Using the bacterial cell suspension prepared in Example 1, a reaction solution containing 1 g / dL of inosine, 5 g / dL of sodium pyrophosphate, and 2 mM of magnesium sulfate in an acetate buffer solution having a pH of 5.0 was prepared. The reaction was carried out at 15-60 ° C. As shown in FIG. 2, each of Escherichia blattae JCM 1650, Klebsiella pneumoniae IFO 3318, Morganella morganii IFO 3168, Enterobacter aerogenes IFO 12010, Chromobacterium fluviatile IAM 13652, and Cedecea lapagei JCM 1684 used in the reaction was 30 to 40 ° C. Serratia ficaria ATCC 33105 produced 5'-inosinic acid most efficiently at around 55 ° C.
[0027]
(Example 6)
Using the strain prepared in Example 1, the reaction was carried out in the same manner using uridine or cytidine instead of inosine in the reaction solution and disodium phenylphosphate as the phosphate donor. The amount of 5′-uridylic acid or 5′-cytidylic acid produced is shown in Table 4.
[0028]
[Table 4]
[0029]
(Example 7)
Using the strain prepared in Example 1, the reaction was similarly carried out using uridine or cytidine instead of inosine in the reaction solution and disodium carbamyl phosphate as the phosphate donor. The amount of 5′-uridylic acid or 5′-cytidylic acid produced is shown in Table 5.
[0030]
[Table 5]
[0031]
【The invention's effect】
A microorganism having an ability to produce a nucleoside-5′-phosphate ester is allowed to act on a nucleoside and a phosphate donor selected from the group consisting of polyphosphoric acid (salt), phenylphosphoric acid (salt) and carbamylphosphoric acid (salt). The method of the present invention for obtaining a -5'-phosphate ester has an effect that a nucleoside-5'-phosphate ester can be produced and collected stably and efficiently without accompanying isomer by-products.
[Brief description of the drawings]
FIG. 1 is a diagram showing the relationship between reaction pH and the amount of 5′-inosinic acid produced.
FIG. 2 is a diagram showing the relationship between the reaction temperature and the amount of 5′-inosinic acid produced.
Claims (3)
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JP31599694A JP3651036B2 (en) | 1993-12-27 | 1994-11-28 | Method for producing nucleoside-5'-phosphate ester |
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JP31599694A JP3651036B2 (en) | 1993-12-27 | 1994-11-28 | Method for producing nucleoside-5'-phosphate ester |
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JPH0937785A (en) * | 1995-05-25 | 1997-02-10 | Ajinomoto Co Inc | Production of nucleoside-5'-phosphate |
JP4304727B2 (en) * | 1996-11-21 | 2009-07-29 | 味の素株式会社 | Method for producing nucleoside-5'-phosphate ester |
US7326546B2 (en) | 2005-03-10 | 2008-02-05 | Ajinomoto Co., Inc. | Purine-derived substance-producing bacterium and a method for producing purine-derived substance |
KR101173533B1 (en) | 2006-04-24 | 2012-08-13 | 아지노모토 가부시키가이샤 | Bacterium capable of producing purine substance, and process for production of purine substance |
JP5104754B2 (en) | 2006-04-24 | 2012-12-19 | 味の素株式会社 | Purine substance producing bacteria and method for producing purine substance |
JP2010110216A (en) | 2007-02-20 | 2010-05-20 | Ajinomoto Co Inc | Method for producing l-amino acid or nucleic acid |
JP2011067095A (en) | 2008-01-10 | 2011-04-07 | Ajinomoto Co Inc | Method for producing target substance by fermentation process |
KR101599977B1 (en) | 2008-02-25 | 2016-03-04 | 아지노모토 가부시키가이샤 | Process for production of 5'-guanylic acid |
JP5636648B2 (en) | 2009-08-10 | 2014-12-10 | 味の素株式会社 | Process for producing 5'-guanylic acid |
WO2015060391A1 (en) | 2013-10-23 | 2015-04-30 | 味の素株式会社 | Method for producing target substance |
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