Rhodobacter capsulatus 유래의 hemA 유전자를 항시발현 벡터(pHCEIIB vector)에 클로닝한 후 과발현시킨 Escherichia coli BLR(DE3) 균주를 사용하여, 5-aminolevulinic acid (ALA) 생산을 위한 적정 배양조건을 조사하였다. ALA 생산을 위한 배양온도는 $37^{circ}C$보다는 $30^{circ}C$에서 더 나은 결과를 나타내었다. Glycine 농도가 세포생장에 미치는 영향이 컸으며, 세포생장을 저해하지 않고ALA를 고농도로 생산하기 위한 적정 glycine 농도는 5-10 g/L 수준이었다. Succinic acid의 적정 첨가수준은 10-20 g/L이었으며, succinic acid의 일부를 glutamate로 대체할 경우에 ALA의 생산량을 향상시키는 효과가 있었다. 한편, 배지 중의 glucose 첨가는 ALA 생산성을 저하시키는 결과를 초래하였다. 이상과 같은 배양 조건 최적화를 통한 jar-fermentor 발효실험을 통하여, 고가의 유도제(IPTG)나 ALA dehydratase inhibitor를 첨가하지 않고도 8.2 g/L 수준의 고농도 ALA를 얻을 수 있었다.
Recombinant Escherichia coli BLR(DE3) harboring the hemA gene from Rhodobacter capsulatus under the control of a constitutive promoter, which we constructed previously, was used for the extracellular production of 5-aminolevulinic acid (ALA). The effects of several factors on ALA production were investigated in flask culture. ALA production by the recombinant E. coli was more efficient at $30^{circ}C$ than $37^{circ}C$. The glycine concentration had an important effect on cell growth. Glycine and succinic acid concentration of 5-10 and 10-20 g/L, respectively, resulted in high ALA production. In addition, the partial replacement of succinic acid by sodium glutamate increased the ALA production. The ALA production was inhibited by the presence of glucose in the medium. Using the optimal conditions, an ALA concentration of 8.2 g/L was achieved in jar fermentation without an added inducer or ALA dehydratase inhibitor; this is the highest reported concentration.
