초록
재조합 인간상피세포 성장인자(rhEGF)가 E. coli BL21(pYHB101) 균주를 사용하여 발현되었다. 10g/L glucose를 첨가한 변형된 MBL 배지를 사용하여 10 $\mu\textrm{m}$ IPTG/lactose로 2시간 동안 유도배양한 후 27$^{\circ}C$에서 48시간 동안 배양하였을 때 44.5 mg/L의 rhEGF가 발현되었다. 상기의 결과는 E. coli BL2l(pYHB101)를 사용하여 rhEGF를 발현시 lactose를 IPTG와 동일한 유도 물질로 사용 가능하다는 것을 시사하는 것이다. 회분식 배양에서 glucose를 10 g/L 첨가한 변형된 MBL 배지에 유도물질로 10 $\mu\textrm{m}$ lactose를 사용하였으며 28시간 동안 배양하였을 때 최대 45 mg/L의 rhEGF가 발현되었다. 유가식 배양에서 정지기에 0.5%(w/v) lactose와 0.25%(w/v) yeast extract를 첨가하였을 때 160mg/L의 rhEGF가 발현되었으며 94.3%가 분비되었다. 이에 비하여 유도기에 lactose를 첨가한 경우는 120 mg/L의 rhEGF가 발현되었으며 cytoplasm으로 발현된 불용성 봉입체의 비율은 20.9%에 달하였다. 이것은 lactose의 첨가시기가 E. coli BL2l(pYHB101)로부터 soluble rhEGF의 생성에 중요하다는 것을 확인한 결과이다.
Recombinant human epidermal growth factor (rhEGF) was produced by E. coli BL2l (DE3) harboring a plasmid pYHB101. The production of rhEGF was 44.5 mg/L when the E. coli BL2l (pYHB101) was cultured at 27$^{\circ}C$ for 48 hr in the modified MBL medium containing 10 $\mu\textrm{g}$/L glucose with 10 $\mu\textrm{m}$ IPTG/lactose induction at 2 hr after inoculation. It was shown that lactose is able to induce the rhEGF expression of E. coli BL2l (pYHB101) with the same efficiency as IPTG. In the batch culture system, when induced with 10 $\mu\textrm{m}$ lactose, E. coli BL2l (pYHB101) produced maximum 45 mg/L of the rhEGF at 28 hr culture in the modified MBL medium containing 10 g/L glucose. In the semi-fed batch culture system, the volumetric yield was 160 mg/L when the culture was added with 0.5% (w/v) lactose and 0.25% (w/v) yeast extract in the late logarithmic phase and 94.3% of rhEGF was secreted as soluble form. However, when the culture was added with them in the early logarithmic phase, the volumetric yield was 120 mg/L and 20.9% of rhEGF was found in cytoplasmic insoluble aggregates. It was found that the addition time of lactose was important for production of soluble rhEGF from E. coli BL21 (pYHB101).
