JPH0561914B2 - - Google Patents
Info
- Publication number
- JPH0561914B2 JPH0561914B2 JP17527085A JP17527085A JPH0561914B2 JP H0561914 B2 JPH0561914 B2 JP H0561914B2 JP 17527085 A JP17527085 A JP 17527085A JP 17527085 A JP17527085 A JP 17527085A JP H0561914 B2 JPH0561914 B2 JP H0561914B2
- Authority
- JP
- Japan
- Prior art keywords
- histidine
- strain
- medium
- strains
- brevibacterium
- 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.)
- Expired - Lifetime
Links
- HNDVDQJCIGZPNO-YFKPBYRVSA-N L-histidine Chemical compound OC(=O)[C@@H](N)CC1=CN=CN1 HNDVDQJCIGZPNO-YFKPBYRVSA-N 0.000 claims description 40
- 229960002885 histidine Drugs 0.000 claims description 22
- 241000186146 Brevibacterium Species 0.000 claims description 9
- 229930001119 polyketide Natural products 0.000 claims description 9
- 125000000830 polyketide group Chemical group 0.000 claims description 7
- 241000186216 Corynebacterium Species 0.000 claims description 6
- 238000004519 manufacturing process Methods 0.000 claims description 6
- HNDVDQJCIGZPNO-UHFFFAOYSA-N histidine Natural products OC(=O)C(N)CC1=CN=CN1 HNDVDQJCIGZPNO-UHFFFAOYSA-N 0.000 claims description 5
- 244000005700 microbiome Species 0.000 claims description 5
- 238000012258 culturing Methods 0.000 claims description 2
- 239000007788 liquid Substances 0.000 claims description 2
- 238000000034 method Methods 0.000 description 10
- HPNSFSBZBAHARI-UHFFFAOYSA-N micophenolic acid Natural products OC1=C(CC=C(C)CCC(O)=O)C(OC)=C(C)C2=C1C(=O)OC2 HPNSFSBZBAHARI-UHFFFAOYSA-N 0.000 description 7
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 6
- 230000001580 bacterial effect Effects 0.000 description 5
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 4
- 210000004027 cell Anatomy 0.000 description 4
- YBJHBAHKTGYVGT-ZKWXMUAHSA-N (+)-Biotin Chemical compound N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 YBJHBAHKTGYVGT-ZKWXMUAHSA-N 0.000 description 3
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- 229920001817 Agar Polymers 0.000 description 3
- 241000186226 Corynebacterium glutamicum Species 0.000 description 3
- 239000008272 agar Substances 0.000 description 3
- 238000000855 fermentation Methods 0.000 description 3
- 230000004151 fermentation Effects 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 231100000350 mutagenesis Toxicity 0.000 description 3
- 230000035772 mutation Effects 0.000 description 3
- 229910052757 nitrogen Inorganic materials 0.000 description 3
- NWUYHJFMYQTDRP-UHFFFAOYSA-N 1,2-bis(ethenyl)benzene;1-ethenyl-2-ethylbenzene;styrene Chemical compound C=CC1=CC=CC=C1.CCC1=CC=CC=C1C=C.C=CC1=CC=CC=C1C=C NWUYHJFMYQTDRP-UHFFFAOYSA-N 0.000 description 2
- MLTVHORGISIISB-UHFFFAOYSA-N 2-hexanoyl-1,3,8-trihydroxy-6-methoxy-anthraquinone Chemical compound COC1=CC(O)=C2C(=O)C3=C(O)C(C(=O)CCCCC)=C(O)C=C3C(=O)C2=C1 MLTVHORGISIISB-UHFFFAOYSA-N 0.000 description 2
- LRFVTYWOQMYALW-UHFFFAOYSA-N 9H-xanthine Chemical compound O=C1NC(=O)NC2=C1NC=N2 LRFVTYWOQMYALW-UHFFFAOYSA-N 0.000 description 2
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- MCPKJGRRWINKOH-UHFFFAOYSA-N Fallacinal Chemical compound C1=C(C=O)C=C2C(=O)C3=CC(OC)=CC(O)=C3C(=O)C2=C1O MCPKJGRRWINKOH-UHFFFAOYSA-N 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
- WHUUTDBJXJRKMK-VKHMYHEASA-N L-glutamic acid Chemical compound OC(=O)[C@@H](N)CCC(O)=O WHUUTDBJXJRKMK-VKHMYHEASA-N 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 229940024606 amino acid Drugs 0.000 description 2
- 150000001413 amino acids Chemical class 0.000 description 2
- QGZKDVFQNNGYKY-UHFFFAOYSA-N ammonia Natural products N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 2
- 239000004202 carbamide Substances 0.000 description 2
- 229910052799 carbon Inorganic materials 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- UYTPUPDQBNUYGX-UHFFFAOYSA-N guanine Chemical compound O=C1NC(N)=NC2=C1N=CN2 UYTPUPDQBNUYGX-UHFFFAOYSA-N 0.000 description 2
- 229910001410 inorganic ion Inorganic materials 0.000 description 2
- 238000002703 mutagenesis Methods 0.000 description 2
- HPNSFSBZBAHARI-RUDMXATFSA-N mycophenolic acid Chemical compound OC1=C(C\C=C(/C)CCC(O)=O)C(OC)=C(C)C2=C1C(=O)OC2 HPNSFSBZBAHARI-RUDMXATFSA-N 0.000 description 2
- 229960000951 mycophenolic acid Drugs 0.000 description 2
- 239000008363 phosphate buffer Substances 0.000 description 2
- 150000003881 polyketide derivatives Chemical class 0.000 description 2
- -1 terrin Chemical compound 0.000 description 2
- 229960000344 thiamine hydrochloride Drugs 0.000 description 2
- 235000019190 thiamine hydrochloride Nutrition 0.000 description 2
- 239000011747 thiamine hydrochloride Substances 0.000 description 2
- DPJRMOMPQZCRJU-UHFFFAOYSA-M thiamine hydrochloride Chemical compound Cl.[Cl-].CC1=C(CCO)SC=[N+]1CC1=CN=C(C)N=C1N DPJRMOMPQZCRJU-UHFFFAOYSA-M 0.000 description 2
- NSYSSMYQPLSPOD-UHFFFAOYSA-N triacetate lactone Chemical compound CC1=CC(O)=CC(=O)O1 NSYSSMYQPLSPOD-UHFFFAOYSA-N 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- GVEZIHKRYBHEFX-MNOVXSKESA-N 13C-Cerulenin Natural products CC=CCC=CCCC(=O)[C@H]1O[C@@H]1C(N)=O GVEZIHKRYBHEFX-MNOVXSKESA-N 0.000 description 1
- YYAYLSOQGBAUHK-UHFFFAOYSA-N 2-(1,3-thiazol-2-ylamino)propanoic acid Chemical compound OC(=O)C(C)NC1=NC=CS1 YYAYLSOQGBAUHK-UHFFFAOYSA-N 0.000 description 1
- USFZMSVCRYTOJT-UHFFFAOYSA-N Ammonium acetate Chemical compound N.CC(O)=O USFZMSVCRYTOJT-UHFFFAOYSA-N 0.000 description 1
- 239000005695 Ammonium acetate Substances 0.000 description 1
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 1
- 239000004475 Arginine Substances 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 241001485655 Corynebacterium glutamicum ATCC 13032 Species 0.000 description 1
- ONIBWKKTOPOVIA-BYPYZUCNSA-N L-Proline Chemical compound OC(=O)[C@@H]1CCCN1 ONIBWKKTOPOVIA-BYPYZUCNSA-N 0.000 description 1
- ODKSFYDXXFIFQN-BYPYZUCNSA-P L-argininium(2+) Chemical compound NC(=[NH2+])NCCC[C@H]([NH3+])C(O)=O ODKSFYDXXFIFQN-BYPYZUCNSA-P 0.000 description 1
- COLNVLDHVKWLRT-QMMMGPOBSA-N L-phenylalanine Chemical compound OC(=O)[C@@H](N)CC1=CC=CC=C1 COLNVLDHVKWLRT-QMMMGPOBSA-N 0.000 description 1
- JLVVSXFLKOJNIY-UHFFFAOYSA-N Magnesium ion Chemical compound [Mg+2] JLVVSXFLKOJNIY-UHFFFAOYSA-N 0.000 description 1
- VZUNGTLZRAYYDE-UHFFFAOYSA-N N-methyl-N'-nitro-N-nitrosoguanidine Chemical compound O=NN(C)C(=N)N[N+]([O-])=O VZUNGTLZRAYYDE-UHFFFAOYSA-N 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Natural products OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 description 1
- 108010073771 Soybean Proteins Proteins 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 1
- JZRWCGZRTZMZEH-UHFFFAOYSA-N Thiamine Natural products CC1=C(CCO)SC=[N+]1CC1=CN=C(C)N=C1N JZRWCGZRTZMZEH-UHFFFAOYSA-N 0.000 description 1
- 241000319304 [Brevibacterium] flavum Species 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 229940043376 ammonium acetate Drugs 0.000 description 1
- 235000019257 ammonium acetate Nutrition 0.000 description 1
- 150000003863 ammonium salts Chemical class 0.000 description 1
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 1
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 1
- 235000011130 ammonium sulphate Nutrition 0.000 description 1
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
- 229960002685 biotin Drugs 0.000 description 1
- 235000020958 biotin Nutrition 0.000 description 1
- 239000011616 biotin Substances 0.000 description 1
- GVEZIHKRYBHEFX-UHFFFAOYSA-N caerulein A Natural products CC=CCC=CCCC(=O)C1OC1C(N)=O GVEZIHKRYBHEFX-UHFFFAOYSA-N 0.000 description 1
- 229910000019 calcium carbonate Inorganic materials 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 235000014633 carbohydrates Nutrition 0.000 description 1
- 239000003729 cation exchange resin Substances 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- GVEZIHKRYBHEFX-NQQPLRFYSA-N cerulenin Chemical compound C\C=C\C\C=C\CCC(=O)[C@H]1O[C@H]1C(N)=O GVEZIHKRYBHEFX-NQQPLRFYSA-N 0.000 description 1
- 229950005984 cerulenin Drugs 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 239000013078 crystal Substances 0.000 description 1
- 210000004748 cultured cell Anatomy 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000002349 favourable effect Effects 0.000 description 1
- RJPAAOHQLUUTRQ-KSVPUCKHSA-N frenolicin Chemical compound O=C1C2=C(O)C=CC=C2C(=O)[C@]23C[C@@H](CC(O)=O)O[C@H](CCC)[C@]12O3 RJPAAOHQLUUTRQ-KSVPUCKHSA-N 0.000 description 1
- HHZQTKVPSKFGTN-UHFFFAOYSA-N frenolicin Natural products CCCC1OC(CC(=O)O)CC23OC12C(=O)c4cc(O)ccc4C3=O HHZQTKVPSKFGTN-UHFFFAOYSA-N 0.000 description 1
- 229960002989 glutamic acid Drugs 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 230000001939 inductive effect Effects 0.000 description 1
- 238000001802 infusion Methods 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 239000003456 ion exchange resin Substances 0.000 description 1
- 229920003303 ion-exchange polymer Polymers 0.000 description 1
- 229910001425 magnesium ion Inorganic materials 0.000 description 1
- 239000013028 medium composition Substances 0.000 description 1
- 239000011785 micronutrient Substances 0.000 description 1
- 235000013369 micronutrients Nutrition 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 235000005985 organic acids Nutrition 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- COLNVLDHVKWLRT-UHFFFAOYSA-N phenylalanine Natural products OC(=O)C(N)CC1=CC=CC=C1 COLNVLDHVKWLRT-UHFFFAOYSA-N 0.000 description 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 1
- 229910001414 potassium ion Inorganic materials 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 239000011347 resin Substances 0.000 description 1
- 229920005989 resin Polymers 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 229910001415 sodium ion Inorganic materials 0.000 description 1
- 235000019710 soybean protein Nutrition 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 239000011593 sulfur Substances 0.000 description 1
- 229910052717 sulfur Inorganic materials 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 235000019157 thiamine Nutrition 0.000 description 1
- 229960003495 thiamine Drugs 0.000 description 1
- 239000011721 thiamine Substances 0.000 description 1
- KYMBYSLLVAOCFI-UHFFFAOYSA-N thiamine Chemical compound CC1=C(CCO)SCN1CC1=CN=C(C)N=C1N KYMBYSLLVAOCFI-UHFFFAOYSA-N 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 229940075420 xanthine Drugs 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Description
〔産業上の利用分野〕
Ｌ−ヒスチジンはアミノ酸輸液及び総合アミノ
酸製剤等の重要な成分である。本発明はこのＬ−
ヒスチジンを発酵法で製造する方法を改良するも
のである。
〔従来技術〕
ブレビバクテリウム属及びコリネバクテリウム
属の微生物がＬ−ヒスチジン生産能を有するため
には、２−チアゾールアラニン（以下２−TAと
略す）米国特許第3716453号）２−TA及び／又
はサルフア剤（特公昭51−23594号公報）等への
耐性を付与せしめれば良いことがわかつている。
更に、２−TA耐性の他にアルギニン、フエニー
ルアラニン、プロリン、キサンチン、グアニン等
を要求する変異株（特公昭51−23593、51−24594
号公報）を使用する方法等が知られている。
〔発明が解決しようとする問題点〕
本発明が解決しようとする問題点はＬ−ヒスチ
ジンの製造コストを低下させるために発酵収率を
向上させることにある。
〔問題点を解決するための手段〕
本発明は上記問題点を解決するためになされた
ものであり、従来より知られているブレビバクテ
リウム属又はコリネバクテリウム属に属するＬ−
ヒスチジン生産能を有する微生物を改良して更に
発酵収率の向上した菌株を見い出すべく研究した
結果、ポリケタイド類に耐性を付与した菌株の中
に、従来のＬ−ヒスチジン生産菌よりも高収率で
Ｌ−ヒスチジンを生産する菌株が存在することを
発明した。
即ち、本発明はブレビバクテリウム属又はコリ
ネバクテリウム属に属し、ポリケタイド類に耐性
を有し、且つＬ−ヒスチジン生産能を有する微生
物を液体培地中で培養し、培地中に生成・蓄積し
たＬ−ヒスチジンを採取することを特徴とするＬ
−ヒスチジンの製造法に関する。
ポリケタイド類としては多くのものが知られて
いるが、例えばtriacetic acid lactone，
mycophenolic acid，terrin，cerulenin，
fallacinal，frenolicin，solorinic acid等がある。
本発明において用いられる微生物はブレビバク
テリウム属又はコリネバクテリウム属に属し、ポ
リケタイド類に耐性を有し、かつＬ−ヒスチジン
生産能を有する変異株である。本発明の変異株を
得るには、下記野生株に、先にＬ−ヒスチジン生
産能を付与し、次いでポリケタイド類耐性を付与
しても良いし、下記野生株にポリケタイド類耐性
を付与し、次いでＬ−ヒスチジン生産能を付与し
ても良い。
本変異株の親株となる野生株は、ブレビバクテ
リウム属又はコリネバクテリウム属の特にコリネ
ホルムＬ−グルタミン酸生産菌として知られてい
るものであり、例えば、以下のものがある。
ブレビバクテリウム・デイバリカタム
ATCC 14022
ブレビバクテリウム・ブラバム ATCC 14067
ブレビバクテリウム・ラクトフエルメンタム
ATCC 13869
ブレビバクテリウム・サツカロリテイカム
ATCC 14066
コリネバクテリウム・アセトアシトフイルム
ATCC 13870
コリネバクテリウム・グルタミクム
ATCC 13032
これらの親株より本発明の変異株を変異誘導す
る方法はＮ−メチル−N′−ニトロ−Ｎ−ニトロ
ソグアニジンに接触せしめる等の通常の変異誘導
方法が適宜適用できる。変異処理した菌株から本
発明の変異株を分離する方法は、ポリケタイド類
を含む培地で生育するような菌株を採取すること
によつて行われる。
本発明の変異株の具体的な変異誘導方法はポリ
ケタイド類の中の一つであるミコフエノール酸
（以下MPAと略す）濃度と菌株の生育度の関係を
以下に示す。
〔変異誘導法〕
ブイヨン寒天スラント上に30℃で24時間生育さ
せたブレビバクテリウム・フラバムAJ11169、
FERM−P4161（ATCC14067より誘導した２−
TA耐性株）及びコリネバクテリウム・グルタミ
クムAJ12092、FERM−P7273（ATCC13032より
誘導した２−TA耐性株）の菌株をＭ／30リン酸
緩衝液に懸濁し菌体濃度108〜109／mlの菌体懸濁
液に500μｇ／mlのＮ−メチル−N′−ニトロ−Ｎ
−ニトロソグアニジンを加え30℃に20分間保持し
た。ついで遠心分離して菌体を集め、Ｍ／30リン
酸緩衝液で良く洗滌した後、下記組成の培地に接
種し31.5℃で４〜10日間培養した。
培地組成（PH7.0）
成分 含量
グリコース 1.0 ｇ／dl
尿 素 0.2 〃
KH2PO4 0.1 〃
MgSO4・7H2Ｏ 0.1 〃
FeSO4・7H2Ｏ 0.002 〃
MnSO4・7H2Ｏ 0.002 〃
ビオチン 100 μｇ／
サイアミン塩酸塩 100 〃
MPA 0.1 ｇ／dl
寒 天 2.0 〃
寒天培地に生育した菌株の中からＬ−ヒスチジ
ン生産能を高い菌株としてブレビバクテリウム・
フラバムAJ12249、FERM−P8371（２−TA耐
性、MPA耐性）及びコリネバクテリウム・グル
タミクムAJ12250、FERM−P8372（２−TA耐
性、MPA耐性）を得た。
このようにして得られた変異株のMPA耐性度
を親株と比較した。
グリコース0.5ｇ／dl、尿素0.15ｇ／dl、硫安
0.15ｇ／dl、KH2PO40.3ｇ／dl、K2HPO40.1
ｇ／dl、MgSO4・7H2O0.01ｇ／dl、CaCl2・2H2
O0.1mg／dl、ビオチン100μｇ／、サイアミン
塩酸塩100μｇ／、FeSO4・7H2O0.002ｇ／dl、
MnSO4・7H2O0.002ｇ／dl及び表に示す量の
MPAを含み、PH7.0に調節した培地に、天然培地
（ペプトン１ｇ／dl、酵母エキス１ｇ／dl、
NaCl0.5ｇ／dl、PH7.0）スラントで24時間培養し
た菌体を殺菌水に懸濁して接触し、24時間培養し
て生育度を濁度で測定した。
[Industrial Application Field] L-histidine is an important component of amino acid infusions, comprehensive amino acid preparations, etc. The present invention is based on this L-
This method improves the method for producing histidine by fermentation. [Prior Art] In order for microorganisms of the genus Brevibacterium and Corynebacterium to have the ability to produce L-histidine, 2-thiazolealanine (hereinafter abbreviated as 2-TA) (US Pat. No. 3,716,453) 2-TA and/or Alternatively, it has been found that it is sufficient to impart resistance to sulfur drugs (Japanese Patent Publication No. 51-23594).
Furthermore, in addition to 2-TA resistance, mutant strains that require arginine, phenylalanine, proline, xanthine, guanine, etc.
A method using the method (No. [Problems to be Solved by the Invention] The problems to be solved by the present invention are to improve the fermentation yield in order to reduce the production cost of L-histidine. [Means for Solving the Problems] The present invention has been made to solve the above-mentioned problems, and is aimed at solving the problems described above.
As a result of research to improve microorganisms capable of producing histidine and find strains with improved fermentation yields, some strains with resistance to polyketides were found to have higher yields than conventional L-histidine producing bacteria. It was discovered that there are strains that produce L-histidine. That is, the present invention cultivates a microorganism belonging to the genus Brevibacterium or Corynebacterium, which is resistant to polyketides and has the ability to produce L-histidine, in a liquid medium, and the L-histidine produced and accumulated in the medium. - L characterized by collecting histidine;
-Relating to a method for producing histidine. Many polyketides are known, such as triacetic acid lactone,
mycophenolic acid, terrin, cerulenin,
There are fallacinal, frenolicin, solorinic acid, etc. The microorganism used in the present invention belongs to the genus Brevibacterium or Corynebacterium, and is a mutant strain that is resistant to polyketides and has the ability to produce L-histidine. To obtain the mutant strain of the present invention, the following wild strain may be first endowed with L-histidine production ability and then polyketide resistance may be imparted, or the following wild strain may be endowed with polyketide resistance and then L-histidine producing ability may be imparted. The wild strain that serves as the parent strain of this mutant strain is a strain of the genus Brevibacterium or Corynebacterium that is particularly known as a coryneform L-glutamic acid producing strain, and includes, for example, the following. Brevibacterium deivalicatum
ATCC 14022 Brevibacterium brabum ATCC 14067 Brevibacterium lactofermentum
ATCC 13869 Brevibacterium satucaroliticum
ATCC 14066 Corynebacterium acetoacitophilum
ATCC 13870 Corynebacterium glutamicum
ATCC 13032 To induce mutagenesis of the mutant strain of the present invention from these parent strains, conventional mutagenesis methods such as contacting with N-methyl-N'-nitro-N-nitrosoguanidine can be appropriately applied. The method for isolating the mutant strain of the present invention from a mutation-treated strain is carried out by collecting a strain that grows in a medium containing polyketides. A specific method for inducing mutations in the mutant strain of the present invention is as follows: The relationship between the concentration of mycophenolic acid (hereinafter abbreviated as MPA), which is one of the polyketides, and the growth rate of the strain is shown below. [Mutation induction method] Brevibacterium flavum AJ11169 grown on broth agar slant at 30°C for 24 hours,
FERM-P4161 (2- derived from ATCC14067
Strains of Corynebacterium glutamicum AJ12092 and FERM-P7273 (2-TA resistant strains derived from ATCC13032) were suspended in M/30 phosphate buffer to a bacterial cell concentration of 10 8 to 10 9 /ml. 500 μg/ml of N-methyl-N'-nitro-N to the bacterial suspension
- Nitrosoguanidine was added and kept at 30°C for 20 minutes. The cells were then collected by centrifugation, washed thoroughly with M/30 phosphate buffer, and then inoculated into a medium having the following composition and cultured at 31.5°C for 4 to 10 days. Medium composition (PH7.0) Ingredient Content Glyose 1.0 g/dl Urea 0.2 〃 KH 2 PO 4 0.1 〃 MgSO 4・7H 2 O 0.1 〃 FeSO 4・7H 2 O 0.002 〃 MnSO 4・7H 2 O 0.002 〃 Biotin 100 μg/thiamine hydrochloride 100 〃 MPA 0.1 g/dl Agar 2.0 〃 Among the strains grown on agar medium, Brevibacterium was selected as a strain with high L-histidine production ability.
flavum AJ12249, FERM-P8371 (2-TA resistant, MPA resistant) and Corynebacterium glutamicum AJ12250, FERM-P8372 (2-TA resistant, MPA resistant). The MPA resistance of the mutant strain thus obtained was compared with that of the parent strain. glycose 0.5g/dl, urea 0.15g/dl, ammonium sulfate
0.15g/dl, KH 2 PO 4 0.3g/dl, K 2 HPO 4 0.1
g/dl, MgSO 4・7H 2 O0.01g/dl, CaCl 2・2H 2
O0.1mg/dl, biotin 100μg/, thiamine hydrochloride 100μg/, FeSO 4・7H 2 O0.002g/dl,
MnSO 4・7H 2 O0.002g/dl and the amount shown in the table
Natural medium (peptone 1g/dl, yeast extract 1g/dl,
(NaCl 0.5 g/dl, PH 7.0) Bacterial cells cultured in a slant for 24 hours were suspended in sterilized water and brought into contact with the cells, cultured for 24 hours, and the degree of growth was measured by turbidity.
【表】【table】
このような変異株を培養する最に用いる培地は
炭素源、窒素源、無機イオン、上記要求性を満足
させるべき物質及び必要に応じビタミン等、その
他の有機微量栄養素を含有する通常の培地であ
る。
炭素源としてはグルコース、シユクロース等の
炭水化物、酢酸等の有機酸等が、窒素源としては
アンモニア水、アンモニアガス、アンモニウム塩
等が好適である。無機イオンとしてはカリイオ
ン、ナトリウムイオン、マグネシウムイオン、リ
ン酸イオン、その他を必要に応じ適宜培地に添加
させる。
培地は好気条件が望ましく、培養の間培地のPH
を４ないし８に温度を25℃ないし37℃に調節しつ
つ行えばより好ましい結果が得られる。かくして
１ないし７日間も培養すれば培地中に著量のＬ−
ヒスチジンが生成蓄積される。
培養液よりＬ−ヒスチジンを採取する方法は、
イオン交換樹脂による方法等通常の方法が採用さ
れる。
以下実施例にて説明する。
実施例
グルコース10ｇ／dl、（NH4）2SO44.5ｇ／dl、
KH2PO40.2ｇ／dl、MgSO4／7H2O0.2ｇ／dl、
FeSO4／7H2O1mg／dl、MnSO4／7H2O1mg／dl、
サイアミン・HCl100μｇ／、ピオチン100μｇ／
、酢酸アンモニウム1.0ｇ／dl、大豆蛋白酸加
水分解液70mg／dl、（全窒素として）、炭酸カルシ
ウム５ｇ／dl（別殺菌）を含む培地をPH7.0に調
節し、その20mlを500ml容肩付フラスコに入れ加
熱殺菌した。これに第１表に示す菌株を一白金耳
接種し、31.5℃に保ちつつ４日間振盪した。各菌
株の培養液中には第２表に示す量のＬ−ヒスチジ
ンがそれぞれ生成蓄積していた。
The medium used for culturing such mutant strains is a normal medium containing carbon sources, nitrogen sources, inorganic ions, substances that should satisfy the above requirements, and other organic micronutrients such as vitamins as necessary. . Preferred carbon sources include carbohydrates such as glucose and sucrose, organic acids such as acetic acid, and preferred nitrogen sources include aqueous ammonia, ammonia gas, and ammonium salts. As inorganic ions, potassium ions, sodium ions, magnesium ions, phosphate ions, and others are appropriately added to the medium as necessary. It is desirable for the medium to be under aerobic conditions, and the PH of the medium should be maintained during cultivation.
More favorable results can be obtained by adjusting the temperature between 25°C and 37°C. Thus, if the culture is continued for 1 to 7 days, a significant amount of L-
Histidine is produced and accumulated. The method for collecting L-histidine from the culture solution is as follows:
A conventional method such as a method using an ion exchange resin is employed. This will be explained below using examples. Example Glucose 10g/dl, (NH 4 ) 2 SO 4 4.5g/dl,
KH 2 PO 4 0.2g/dl, MgSO 4 /7H 2 O0.2g/dl,
FeSO 4 /7H 2 O1mg/dl, MnSO 4 /7H 2 O1mg/dl,
Thiamine/HCl 100μg/, Piotine 100μg/
A medium containing 1.0 g/dl of ammonium acetate, 70 mg/dl of soybean protein acid hydrolyzate (as total nitrogen), and 5 g/dl of calcium carbonate (separately sterilized) was adjusted to pH 7.0, and 20 ml of it was mixed into a 500 ml volume. The mixture was placed in a flask and sterilized by heating. A loopful of the bacterial strains shown in Table 1 was inoculated into this, and the mixture was shaken for 4 days while being kept at 31.5°C. In the culture solution of each strain, L-histidine was produced and accumulated in the amounts shown in Table 2.
【表】
AJ12249を上記の方法で培養して培養液１を
得、これより遠心分離にて菌体他を除き、上清を
強酸性陽イオン交換樹脂“ダイヤイオンSK−
1B”（NH4 +型）に通過させた。樹脂を水洗後、
2N・NH4OHにてＬ−ヒスチジンを溶出し、つ
いで溶出液を濃縮し、これよりＬ−ヒスチジンの
粗結晶9.8ｇを得た。[Table] AJ12249 was cultured using the above method to obtain culture solution 1, which was centrifuged to remove bacterial cells and the supernatant was collected using a strongly acidic cation exchange resin "Diaion SK-".
1B” (NH 4 + type). After washing the resin with water,
L-histidine was eluted with 2N.NH 4 OH, and the eluate was then concentrated to obtain 9.8 g of crude crystals of L-histidine.
Claims (1)
ム属に属しポリケタイド類に耐性を有し、且つＬ
−ヒスチジン生産能を有する微生物を液体培地中
で培養し、培地中に生成蓄積したＬ−ヒスチジン
を採取する事を特徴とするＬ−ヒスチジンの製造
法。1 Belongs to the genus Brevibacterium or Corynebacterium and is resistant to polyketides, and
- A method for producing L-histidine, which comprises culturing a microorganism capable of producing histidine in a liquid medium and collecting L-histidine produced and accumulated in the medium.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP17527085A JPS6236197A (en) | 1985-08-09 | 1985-08-09 | Production of l-histidine by fermentation method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP17527085A JPS6236197A (en) | 1985-08-09 | 1985-08-09 | Production of l-histidine by fermentation method |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPS6236197A JPS6236197A (en) | 1987-02-17 |
| JPH0561914B2 true JPH0561914B2 (en) | 1993-09-07 |
Family
ID=15993210
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP17527085A Granted JPS6236197A (en) | 1985-08-09 | 1985-08-09 | Production of l-histidine by fermentation method |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPS6236197A (en) |
Families Citing this family (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5188948A (en) * | 1987-04-16 | 1993-02-23 | Ajinomoto Co., Inc. | Process for producing L-valine by fermentation |
| JPH0665314B2 (en) * | 1987-04-16 | 1994-08-24 | 味の素株式会社 | Fermentation method for producing L-valine |
| RU2282660C2 (en) * | 2003-11-10 | 2006-08-27 | Закрытое акционерное общество "Научно-исследовательский институт Аджиномото-Генетика" (ЗАО АГРИ) | Mutant phosphoribozyl pyrophosphate synthetase, dna fragment, bacteria of genus escherichia as l-histidine and method for production of l-histidine |
| US20060039955A1 (en) * | 2004-05-28 | 2006-02-23 | Cargill, Incorporated | Animal feed compositions with enhanced histidine content |
-
1985
- 1985-08-09 JP JP17527085A patent/JPS6236197A/en active Granted
Also Published As
| Publication number | Publication date |
|---|---|
| JPS6236197A (en) | 1987-02-17 |
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