JP2012530515A - L−リシン生産能が向上した微生物及びこれを用いたl−リシン生産方法 - Google Patents
L−リシン生産能が向上した微生物及びこれを用いたl−リシン生産方法 Download PDFInfo
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- JP2012530515A JP2012530515A JP2012524667A JP2012524667A JP2012530515A JP 2012530515 A JP2012530515 A JP 2012530515A JP 2012524667 A JP2012524667 A JP 2012524667A JP 2012524667 A JP2012524667 A JP 2012524667A JP 2012530515 A JP2012530515 A JP 2012530515A
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- Prior art keywords
- lysine
- activity
- microorganism
- producing
- gluconate kinase
- 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.)
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- 239000004472 Lysine Substances 0.000 title claims abstract description 61
- 235000019766 L-Lysine Nutrition 0.000 title claims abstract description 50
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- 238000004519 manufacturing process Methods 0.000 title claims abstract description 34
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- 108010021382 Gluconokinase Proteins 0.000 claims abstract description 60
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- 108090000623 proteins and genes Proteins 0.000 claims description 53
- 241000186226 Corynebacterium glutamicum Species 0.000 claims description 25
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- 238000000926 separation method Methods 0.000 description 1
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- 239000000243 solution Substances 0.000 description 1
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- 239000004455 soybean meal Substances 0.000 description 1
- 239000003549 soybean oil Substances 0.000 description 1
- 235000012424 soybean oil Nutrition 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000008117 stearic acid Substances 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 235000000346 sugar Nutrition 0.000 description 1
- 150000008163 sugars Chemical class 0.000 description 1
- 235000020238 sunflower seed Nutrition 0.000 description 1
- 239000013589 supplement Substances 0.000 description 1
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 1
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Abstract
【選択図】なし
Description
破裂のような遺伝子変異によってGntK活性が下向調節され、よって、天然型微生物での酵素の内在的活性より低くなることを意味する。本発明の一例において、本発明は図2または図3の開裂地図で表される組換えベクターの導入を介してGntK活性を妨害することにより増加されたL−リシン生産能を有するコリネバクテリウム属の微生物を提供する。
組換え菌株KFCC10881−ΔNCgl2399とKFCC10881−ΔNCgl2905の製造に使用するためにプライマーをデザインした。まず、アメリカ国立保健院の遺伝子銀行(NIH Genbank)からNCgl2399(配列番号1)及びNCgl2905(配列番号2)の配列を確保した。前記配列などに基づいてNCgl2399またはNCgl2905の不活性化断片を構成するのに使用されるプライマー2399F1、2399F2、2399R1、2399R2、2905F1、2905F2、2905R1、及び2905R2を合成した(表1)。本発明で使用されたプライマーなどの塩基配列を配列番号と共に表1に要約した。制限サイトに下線を引いた。
組換え菌株KFCC10881−ΔNCgl2399を電気穿孔法を使用して組換えプラスミドpDZ−ΔNCgl2905と形質転換させ、前記示したものと同様な方法で処理してNCgl2399及びNCgl2905遺伝子の両方に欠損された新規な組換え菌株を得た。前記変異菌株をコリネバクテリウム・グルタミクムCA01−0892と命名し、2010年6月24日付けで受託番号KCCM11085Pで韓国微生物培養センターに寄託した。
前記実施例1と2で製作されたL−リシン生産菌株であるコリネバクテリウム・グルタミクムKFCC−10881−ΔNCgl2399、KFCC−10881−ΔNCgl2905及びCA01−0892(KFCC−10881−ΔNCgl2399ΔNCgl2905)の細胞内グルコン酸キナーゼ(GntK)活性及びNADPH水準を下記のような方法で分析した。
グルコース20g、ペプトン10g、酵母エキス5g、尿素1.5g、KH2PO44g、K2HPO48g、MgSO47H2O0.5g、ビオチン100μg、チアミンHCl1000μg、カルシウム−パントテン酸2000μg、ニコチンアミド2000μg(蒸溜水1リットル基準)
L−リシン生産のために、前記実施例1と2で製作されたL−リシン生産菌株であるコリネバクテリウム・グルタミクムKFCC−10881−ΔNCgl2399、KFCC−10881−ΔNCgl2905及びCA01−0892(KFCC−10881−ΔNCgl2399ΔNCgl2905)を次のように培養した。
グルコース20g、ペプトン10g、酵母エキス5g、尿素1.5g、KH2PO44g、K2HPO48g、MgSO47H2O0.5g、ビオチン100μg、チアミンHCl1000μg、カルシウム−パントテン酸2000μg、ニコチンアミド2000μg(蒸溜水1リットル基準)
グルコース100g、(NH4)2SO440g、大豆タンパク質2.5g、トウモロコシ浸漬固形粉5g、尿素3g、KH2PO41g、MgSO47H2O0.5g、ビオチン100μg、チアミンHCl1000μg、カルシウム−パントテン酸2000μg、ニコチンアミド3000μg、CaCO330g(蒸溜水1リットル基準)
Claims (10)
- 内在的活性に比べて弱化されたグルコン酸キナーゼ(GntK)活性を有する、L−リシン生産微生物。
- 前記グルコン酸キナーゼは、配列番号1または配列番号2のアミノ酸配列を有するものである、請求項1に記載のL−リシン生産微生物。
- 前記微生物は、コリネバクテリウム属である、請求項1に記載のL−リシン生産微生物。
- グルコン酸キナーゼをエンコードする染色体遺伝子上の塩基配列全体または一部の欠損、一部の置換または挿入によって内在的グルコン酸キナーゼ活性が弱化されたものである、請求項1に記載のL−リシン生産微生物。
- グルコン酸キナーゼをエンコードする染色体遺伝子の調節要素上で、全体または一部の欠損、一部の置換または挿入によって内在的グルコン酸キナーゼ活性が弱化されたものである、請求項1に記載のL−リシン生産微生物。
- グルコン酸キナーゼ活性を有し、互いに異なるアミノ酸配列を有するタンパク質をコード化する2つまたはそれ以上の染色体遺伝子が存在する場合、少なくとも1つの染色体遺伝子上の全体または一部の欠損、置換または挿入によって;または少なくとも1つの染色体遺伝子の調節要素上で全体または一部が欠損、一部が置換または挿入によって内在的グルコン酸キナーゼ活性が弱化されたものである、請求項1に記載のL−リシン生産微生物。
- 前記微生物は、コリネバクテリウム・グルタミクムKFCC10881から由来したものである、請求項1に記載のL−リシン生産微生物。
- 前記微生物は、受託番号KCCM11085Pで寄託されたコリネバクテリウム・グルタミクムCA01−0892として識別されるものである、請求項1に記載のL−リシン生産微生物。
- 1)全体または一部の配列が変異されて弱化された活性を有するグルコン酸キナーゼ(GntK)をエンコードするポリヌクレオチド断片を構成する段階;
2)組換えベクターを提供するために、宿主細胞で染色体と相同組換えできるベクターに前記収得したポリヌクレオチド断片を挿入する段階;
3)相同組換え体を形成するために、前記収得した組換えベクターをL−リシンを生産することができる宿主細胞へ導入する段階;及び
4)前記相同組換え体の中からGntKの活性が内在的活性に比べて弱化された菌株を選抜する段階を含む、請求項1乃至8のうち、いずれか1項によるL−リシン生産微生物の製造方法。 - 1)請求項1乃至8のうち、いずれか1項による微生物を培養して細胞培養物を収得する段階:及び
2)前記細胞培養物または微生物からL−リシンを回収する段階を含む、L−リシンの生産方法。
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KR10-2010-0068618 | 2010-07-15 | ||
KR20100068618 | 2010-07-15 | ||
KR1020110065694A KR101269810B1 (ko) | 2010-07-15 | 2011-07-01 | L-라이신 생산능이 향상된 미생물 및 이를 이용한 l-라이신 생산방법 |
KR10-2011-0065694 | 2011-07-01 | ||
PCT/KR2011/005252 WO2012008810A2 (en) | 2010-07-15 | 2011-07-15 | Microorganism with enhanced l-lysine productivity and method for producing l-lysine using the same |
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KR (1) | KR101269810B1 (ja) |
CN (1) | CN102741393B (ja) |
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Cited By (2)
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JP2018512877A (ja) * | 2015-04-20 | 2018-05-24 | シージェイ チェイルジェダン コーポレーション | グルコネートリプレッサー変異体、これを含むl−リシンを生産する微生物及びこれを利用したl−リシン生産方法 |
JP2018518977A (ja) * | 2015-07-03 | 2018-07-19 | シージェイ チェイルジェダン コーポレーション | L−リジンを生産する微生物及びそれを用いたl−リジン生産方法 |
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KR101582008B1 (ko) | 2013-10-15 | 2015-12-31 | 씨제이제일제당 (주) | 생물막 형성 억제 활성을 가지는 유전자 및 이 유전자가 불활성화된 균주를 이용한 l-라이신 생산 방법 |
KR101766964B1 (ko) * | 2015-08-27 | 2017-08-09 | 씨제이제일제당 (주) | L-라이신 생산능을 가지는 코리네박테리움 속 미생물 및 이를 이용한 l-라이신 생산방법 |
KR102011394B1 (ko) * | 2017-07-19 | 2019-10-22 | 씨제이제일제당 주식회사 | 퓨트레신을 생산하는 미생물 및 이를 이용한 퓨트레신 생산방법 |
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Publication number | Priority date | Publication date | Assignee | Title |
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JP2018512877A (ja) * | 2015-04-20 | 2018-05-24 | シージェイ チェイルジェダン コーポレーション | グルコネートリプレッサー変異体、これを含むl−リシンを生産する微生物及びこれを利用したl−リシン生産方法 |
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PL2430152T3 (pl) | 2017-02-28 |
ES2603128T3 (es) | 2017-02-23 |
BRPI1106092A2 (pt) | 2016-10-11 |
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CN102741393A (zh) | 2012-10-17 |
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