CAJ | 학술논문

采用基因工程的方法构建重组大肠杆菌得到重组菌Eco（pETDuet-ATA117），实现了转氨酶ATA117的重组表达。使用响应面法对重组大肠杆菌摇瓶发酵产酶培养基进行了优化并对转氨酶诱导条件进行研究。首先采用二水平部分因子设计筛选确定培养基中对转氨酶ATA117酶活有显著影响的3种培养基组分，即甘油、酵母粉和胰蛋白胨。然后进行最陡爬坡实验确定最佳响应面区域，最后通过Box-Behnken设计确定各成分的最佳浓度。采用该优化培养基，转氨酶酶活为391.7 U/L，分别是基础培养基和单因素优化培养基酶活的3.13倍和1.22倍。对转氨酶的诱导条件进行了研究，得出最适诱导温度24℃，光密度值（OD）为0.8，此条件下进行诱导，异丙基硫代半乳糖苷（IPTG）浓度1 mmol/L，诱导时间15 h，最终转氨酶活力达到713.7 U/L。
채용기인공정적방법구건중조대장간균득도중조균Eco（pETDuet-ATA117），실현료전안매ATA117적중조표체。사용향응면법대중조대장간균요병발효산매배양기진행료우화병대전안매유도조건진행연구。수선채용이수평부분인자설계사선학정배양기중대전안매ATA117매활유현저영향적3충배양기조분，즉감유、효모분화이단백동。연후진행최두파파실험학정최가향응면구역，최후통과Box-Behnken설계학정각성분적최가농도。채용해우화배양기，전안매매활위391.7 U/L，분별시기출배양기화단인소우화배양기매활적3.13배화1.22배。대전안매적유도조건진행료연구，득출최괄유도온도24℃，광밀도치（OD）위0.8，차조건하진행유도，이병기류대반유당감（IPTG）농도1 mmol/L，유도시간15 h，최종전안매활력체도713.7 U/L。
The recombinant Eco(pETDuet-ATA117) was constructed for the functional expression of transaminase ATA117. After that, the response surface methodology was employed to optimize the fermentation conditions. A two-level fractional factor design was used to evaluate the effects of different components of medium. The results showed that glycerin, yeast extract and tryptone had important effects on transaminase activity, then the optimal response surface was determined by the Steepest ascent design, finally, the optimal medium composition was observed by Box-Behnken design: glycerin14.45 g/L, yeast extract 7.27 g/L and tryptone 5.72 g/L. Under the optimal conditions, the Enzyme activity(391.7 U/L) was much higher than that using either basal fermentation medium(125.4 U/L) or single variable optimization of fermentation medium(320.7 U/L). Meanwhile, the inducing conditions was also studied, and under the optimal induction conditions of 24℃, optical densityof 0.8 under 600 nm, the final Enzyme activity was 713.7 U/Lafter 15 h induction time when isopropyl thiogalactoside was 1 mmol/L.