메탄올로부터 Methylobacterium organophilum에 의한 Poly-$\beta$-hydroxybutyric Acid의 생산과 배지성분의 최적화

Optimization of Growth Medium and Poly-$\beta$-hydroxybutyric Acid Production from Methanol in Methylobacterium organophilum

  • Choi, Joon-H (Department of Biological Science and Engineering, KAIST) ;
  • Kim, Jung H. (Department of Biological Science and Engineering, KAIST) ;
  • M. Daniel (University de Technologie de Compiegne, Genie Chimique, France) ;
  • J.M. Lebeault (University de Technologie de Compiegne, Genie Chimique, France)
  • 발행 : 1989.08.01

초록

Facultative methylotroph인 Methylobacterium organophilum을 유일한 탄소원으로 메탄올을 0.5% (v/v) 함유한 최소 합성배지를 이용하여 pH 6.8과 3$0^{\circ}C$에서 배양하였다. 세포의 증식은 배지 내의 여러 성분들에 의해 영향을 받았으며 이로부터 최대증식속도를 얻을 수 있는 최적 합성배지 조성을 확립하였다. 배지조성 중 질소원이 결핍되면 세포의 증식이 감소하며 Fe$^{+2}$ 또는 Mn$^{+2}$ 이온의 결핍은 세포의 비증식속도를 감소시켰다. 탄소원인 메탄올은 농도를 증가시킬수록 세포의 비증식속도가 감소하는 메탄올의 기질 저해성을 확인하였으며, 4%(v/v) 이상의 메탄을 농도에서는 세포의 증식이 완전히 저해되었다. 이러한 메탄올의 기질 저해성은 간헐식 유가배양법을 이용하여 극복하고 PHB의 축적을 촉진하는 영양분 제한(nutrient-deficiency) 실험을 수행하였다. 합성배지의 영양분 중 NH$_{4}^{+}$, SO$_{4}^{-2}$, $K^+$ 또는 PO$_{4}^{-3}$ 이온을 결핍시켰을 때 M. organophilum의 PHB 생산이 증가하였으며 칼륨이온을 결핍시켰을때 세포의 건조 중량에 대한 PHB 함량은 최대 58%를 얻었다.

Methylobacterium organophilum, a facultative methylotroph was cultivated on a methanol as a sole carbon and energy source. The cell growth was affected by the various components of minimal synthetic medium and the medium composition was optimized with 0.5% (v/v) methanol at pH 6.8 and at 3$0^{\circ}C$. The maximum specific growth rate of M. organophilum was achieved to 0.26 hr$^{-1}$ in the optimized medium which has following composition: Methanol, 0.5% (v/v):(NH$_4$)$_2$SO$_4$, 1.0g/l:KH$_2$PO$_4$, 2.13g/l:KH$_2$PO$_4$, 1.305g/ι:MgSO$_4$.7$H_2O$. 45g/l and trace elements (CaCl$_2$.2$H_2O$, 3.3mg:FeSO$_4$.7$H_2O$, 1.3mg:MnSO$_4$.4$H_2O$, 130$\mu\textrm{g}$:ZnSO$_4$.5$H_2O$, 40$\mu\textrm{g}$:Na$_2$MoO$_4$.2$H_2O$, 40$\mu\textrm{g}$:CoCl$_2$.6$H_2O$, 40$\mu\textrm{g}$:H$_3$BO$_3$, 30$\mu\textrm{g}$ per liter). By the limitation of nitrogen and deficiency of Mn$^{+2}$ or Fe$^{+2}$, the cell growth was significantly repressed. Methanol greatly repressed the cell growth and the complete inhibition was observed at concentration above 4% (v/v). In order to overcome the methanol inhibition and to prevent the methanol limitation, intermittent feeding of methanol was conducted by a D.O.-stat technique. PHB production by M. organophilum was stimulated by deficiency of nutrients such as NH$_{4}^{+}$, SO$_{4}^{-2}$, $Mg^{+2}$, $K^{+}$, or PO$_{4}^{-3}$ in the medium. The maximum PHB content was obtained as 58% of dry cell weight under deficiency of potassium ion in the optimized synthetic medium.

키워드