JP7475408B2 - L-アルギニンまたはl-シトルリン生産能が向上したコリネバクテリウム属微生物およびこれを用いたl-アルギニンまたはl-シトルリンの生産方法 - Google Patents
L-アルギニンまたはl-シトルリン生産能が向上したコリネバクテリウム属微生物およびこれを用いたl-アルギニンまたはl-シトルリンの生産方法 Download PDFInfo
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- JP7475408B2 JP7475408B2 JP2022162571A JP2022162571A JP7475408B2 JP 7475408 B2 JP7475408 B2 JP 7475408B2 JP 2022162571 A JP2022162571 A JP 2022162571A JP 2022162571 A JP2022162571 A JP 2022162571A JP 7475408 B2 JP7475408 B2 JP 7475408B2
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- arginine
- corynebacterium
- citrulline
- strain
- amino acid
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Description
ynebacterium imitans)、コリネバクテリウムカスピウム(Corynebacterium caspium)、コリネバクテリウムテスツディノリス(Corynebacterium testudinoris)、コリネバクテリウムシュードペラルギ(Corynebacaterium pseudopelargi)またはコリネバクテリウムフラベセンス(Corynebacterium flavescens)であってもよいが、これに限定されるものではない。
本発明で使われた「一部」は、塩基配列またはポリヌクレオチド、またはアミノ酸配列の全部ではないものを意味し、1~300個、好ましくは1~100個、より好ましくは1~50個であってもよいが、これに限定されるものではない。
アセチルオルニチンアミノトランスフェラーゼの活性が強化されたコリネバクテリウムグルタミカム変異株を製造するために、L-アルギニン生産菌株であるコリネバクテリウムグルタミカム14GR(KCCM13219P)菌株およびE.coli DH5a(HIT Competent cellsTM、Cat No.RH618)を使用した。
菌株に生合成経路を強化させるためにアセチルオルニチンアミノトランスフェラーゼに変異を誘導した。本実施例で用いた方法は、アセチルオルニチンアミノトランスフェラーゼを暗号化しているargD遺伝子の発現を増加するために、当該遺伝子の特定位置に変異を誘発した。argD遺伝子の131番目のアミノ酸をアラニンからトレオニンに置換し、コリネバクテリウムグルタミカムゲノム上でargD遺伝子の真ん中を中心に左アーム529bp部分と右アーム525bp部分をPCRで増幅して、オーバーラップPCR(overlap PCR)方法で連結した後、pk19msbベクターにクローニングした。前記プラスミドをpk19msb+argD(A131T)と名付けた(図1参照)。前記プラスミドを作製するためにそれぞれの遺伝子断片を増幅するのに下記表1のプライマーを使用した。
クローニングベクターを用いて、変異菌株であるAD1菌株を製造した。クローニングベクターの最終濃度が1μg/μl以上となるように用意して、コリネバクテリウムグルタミカム14GR菌株に電気穿孔法(文献[Tauch et al.、FEMS Microbiology letters123(1994)343-347]参照)を用いて1次組換えを誘導した。この時、電気穿孔した菌株をカナマイシンが50μg/μl含まれる寒天培地に塗抹してコロニーを分離した後、ゲノム上の誘導した位置に適切に挿入されたかを、PCRおよび塩基配列分析により確認した。このように分離された菌株を再度2次組換えを誘導するために液体培地に接種し、一晩以上培養して10%sucroseを含有した寒天培地に塗抹してコロニーを分離した。最終分離したコロニー中でカナマイシンに対する耐性の有無を確認した後、抗生剤耐性のない菌株のうちアセチルオルニチンアミノトランスフェラーゼに変異が導入したかを、塩基配列分析により確認した(文献[Schafer et al.、Gene145(1994)69-73]参照)。最終的に、アセチルオルニチンアミノトランスフェラーゼのアミノ酸配列(配列番号2)のうち131番目に位置したアラニンがトレオニンに置換された、L-アルギニンの生産が可能なコリネバクテリウムグルタミカム変異株AD1を作製した。
親菌株と比較して、実施例1で作製されたコリネバクテリウムグルタミカム変異株AD1のL-アルギニン生産能を評価した。
実施例1-1のpk19msb+argD(A131T)クローニングベクターを、コリネバクテリウムグルタミカム14GRの代わりに、コリネバクテリウムグルタミカム15GD(KCCM13220P)に導入したことを除けば、実施例1と同様の方法でアセチルオルニチンアミノトランスフェラーゼのアミノ酸配列(配列番号2)のうち131番目に位置したアラニンがトレオニンに置換された、L-シトルリンの生産が可能なコリネバクテリウムグルタミカム変異株CD1を作製した。
親菌株と比較して、実施例2で作製されたコリネバクテリウムグルタミカム変異株CD1のL-シトルリン生産能を評価した。
寄託機関名:韓国微生物保存センター(KCCM)
受託番号:KCCM13219P
受託日付:20220629
寄託機関名:韓国微生物保存センター(KCCM)
受託番号:KCCM13220P
受託日付:20220629
Claims (2)
- 配列番号2のアミノ酸配列のうち、131番目のアミノ酸においてアラニンからトレオニンへの置換を有するアセチルオルニチンアミノトランスフェラーゼを発現することで、親株と比べてL-アルギニンまたはL-シトルリンの生産能が向上したコリネバクテリウム グルタミカム(Corynebacterium glutamicum)変異株。
- 請求項1に記載のコリネバクテリウム グルタミカム変異株を培地で培養するステップと、
前記培養された変異株または変異株が培養された培地からL-アルギニンまたはL-シトルリンを回収するステップとを含むL-アルギニンまたはL-シトルリンの生産方法。
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JP2017079705A (ja) | 2015-10-30 | 2017-05-18 | 味の素株式会社 | グルタミン酸系l−アミノ酸の製造法 |
JP2020523989A (ja) | 2017-06-14 | 2020-08-13 | シージェイ チェイルジェダン コーポレーション | 新規なポリペプチド及びそれを用いたオルニチン系産物の生産方法 |
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KR100854234B1 (ko) * | 2006-07-13 | 2008-08-25 | 씨제이제일제당 (주) | 역가가 향상된 유전자 argF 염기서열 및 그를 포함하는형질전환 균주를 이용한 L―알지닌의 생산방법 |
KR100830289B1 (ko) * | 2007-01-18 | 2008-05-16 | 씨제이제일제당 (주) | L-아르기닌 생산 변이주 및 이의 제조방법 |
KR101348461B1 (ko) * | 2010-12-08 | 2014-01-08 | 씨제이제일제당 (주) | 퓨트레신을 생산하는 미생물 및 이를 이용하여 퓨트레신을 생산하는 방법 |
KR102269637B1 (ko) | 2018-12-26 | 2021-06-28 | 대상 주식회사 | L-시트룰린 또는 l-아르기닌 생산능이 향상된 변이 균주 및 이를 이용한 l-시트룰린 또는 l-아르기닌의 제조 방법 |
-
2022
- 2022-07-21 KR KR1020220090641A patent/KR20240013960A/ko unknown
- 2022-08-17 WO PCT/KR2022/012253 patent/WO2024019215A1/ko unknown
- 2022-10-04 US US17/937,833 patent/US20240026283A1/en active Pending
- 2022-10-06 EP EP22200149.7A patent/EP4310179A1/en active Pending
- 2022-10-07 JP JP2022162571A patent/JP7475408B2/ja active Active
- 2022-10-18 CN CN202211274517.0A patent/CN117431197A/zh active Pending
Patent Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2017079705A (ja) | 2015-10-30 | 2017-05-18 | 味の素株式会社 | グルタミン酸系l−アミノ酸の製造法 |
JP2020523989A (ja) | 2017-06-14 | 2020-08-13 | シージェイ チェイルジェダン コーポレーション | 新規なポリペプチド及びそれを用いたオルニチン系産物の生産方法 |
CN113736719A (zh) | 2020-05-29 | 2021-12-03 | 陕西鸿道生物分析科学技术研究院有限公司 | 一种谷氨酸棒杆菌基因工程菌及其在亚精胺生产中的应用 |
WO2022008280A1 (en) | 2020-07-09 | 2022-01-13 | Evonik Operations Gmbh | Method for the fermentative production of guanidinoacetic acid |
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US20240026283A1 (en) | 2024-01-25 |
EP4310179A1 (en) | 2024-01-24 |
JP2024014656A (ja) | 2024-02-01 |
CN117431197A (zh) | 2024-01-23 |
KR20240013960A (ko) | 2024-01-31 |
WO2024019215A1 (ko) | 2024-01-25 |
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