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Bacillus clausii I-52의 형질 전환에 의한 Alkaline Protease의 생산성 향상 및 Pilot-scale 생산 연구 KCI 등재
Study on the Increased Production of an Alkaline Protease from Bacillus clausii I-52 by Transformation and Pilot-scale Production by Submerged Fermentation
pp.187-196
BCAP 유전자를 지니는 pRB374-BCAP2와 pLip-BCAP2를 B. clausii I-52에 도입 후, 염색체 integration에 의해 구성된 pHPS9-BCAP 형질전환체(B. clausii C5)와 alkaline protease 발현율을 비교하였다. 최적화 배지(대두박 2%, 밀가루 1%, 구연산나트륨 0.5%, K2HPO4 0.4%, Na2HPO4 0.1%, NaCl 0.4%, MgSO4⋅7H2O 0.01%, FeSO4⋅7H2O 0.05%, 물엿 2.5%, 탄산나트륨 0.6%)에서 액침배양(37℃, 48 h, 650 rpm, 1 vvm) 시, pRB374- BCAP2 및 pHPS9-BCAP 형질전환체 각각은 15% 및 61% 정도 alkaline protease 생산이 증가하였다. 그러나 pLip-BCAP2 형질전환체에서는 변화가 없었다. 최고의 활성 균주인, B. clausii C5를 300 L 규모 pilot-scale 액침 배양(37℃, 30 h, 250 rpm, 1 vvm) 시, alkaline protease 생산은 105,685 U/mL로 측정되었다.
Plasmid types, pRB374-BCAP2 and pLip-BCAP2 carrying B. clausii alkaline protease (BCAP) were constructed, introduced into B. clausii I-52, and protease productivities were compared with B. clausii C5 (pHPS9-BCAP), which the BCAP gene was integrated into the chromosomal DNA. The protease yield was increased by 15% for B. clausii R6 (pRB374-BCAP2) and 61% for B. clausii C5 (pHPS9-BCAP) in an optimized medium (soybean meal 2%, wheat flour 1%, sodium citrate 0.5%, K2HPO4 0.4%, Na2HPO4 0.1%, NaCl 0.4%, MgSO4⋅7H2O 0.01%, FeSO4⋅7H2O 0.05%, liquid maltose 2.5%, Na2CO3 0.6%) under the fixed culture condition (37℃, 48 h, 1 vvm, 650 rpm). The B. clausii L7 (pLip-BCAP2) transformant showed no significant difference. B. clausii C5 showing the highest protease yield could produce 105,685 U/mL from 300 L pilot-scale fermentation (37℃, 30 h, 1 vvm, 250 rpm).
