Journal of Life Science (생명과학회지)

Korean Society of Life Science (한국생명과학회)
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Growth Characteristics of a Pyruvate Decarboxylase Mutant Strain of Zymomonas mobilis

Pyruvate decarboxylase 돌연변이 Zymomonas mobilis 균주의 생장 특성 연구

  • Xun, Zhao (School of Biotechnology and Biomolecular Sciences, The University of New South Wales) ;
  • Peter L., Rogers (School of Biotechnology and Biomolecular Sciences, The University of New South Wales) ;
  • Kwon, Eilhann E. (Department of Environment and Energy at Sejong University) ;
  • Jeong, Sang Chul (Freshwater Bioresources Utilization Division, Nakdonggang National Institute of Biological Resources) ;
  • Jeon, Young Jae (School of Biotechnology and Biomolecular Sciences, The University of New South Wales)
  • 순 자오 (호주 뉴사우스 웨일즈 대학 생물분자 생명공학부) ;
  • 피터 로저스 (호주 뉴사우스 웨일즈 대학 생물분자 생명공학부) ;
  • 권일한 (세종대학교 환경에너지융합학과) ;
  • 정상철 (국립낙동강생물자원관 담수 생물특성연구실) ;
  • 전용재 (호주 뉴사우스 웨일즈 대학 생물분자 생명공학부)
  • Received : 2015.08.19
  • Accepted : 2015.10.13
  • Published : 2015.11.30
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Abstract

Studies of the inactivation of a gene encoding pyruvate decarboxylase, pdc, in an ethanol-producing bacterium, Zymomonas mobilis, identified a mutant strain with 50% reduced PDC activity. To evaluate the possibility of a carbon-flux shift from an ethanol pathway toward higher value fermentation products, including pyruvate, succinate, and lactate, fermentation studies were carried out. Despite attempts to silence pdc expression in the wild-type strain ZM4 using cat-inserted pdc and pdc-deleted homologs by electroporation, the strain isolated showed partial gene activation. Fermentation experiments with the PDC mutant strain showed that the reduced expression level of PDC activity resulted in decreased rates of substrate uptake and ethanol production, together with increased pyruvate accumulation of 2.5 g l-1 , although lactate and succinate concentrations were not significantly enhanced in these modified strains. Despite numerous attempts, no strains were isolated in which complete pdc inactivation occurred. This result indicates that the ethanol fermentation pathway of this bacterium is totally dependent on the activity of the PDC enzyme. To ensure a redox balance of intracellular NAD and NADH levels, other enzymes, such as lactate dehydrogenase for lactate, and enzymes involved in the production of succinic acid, such as pyruvate dehydrogenase (PDH) and malic enzymes, may be needed for their increased end-product production.

에탄올 생산 세균 Zymomonas mobilis에서 에탄올 생산 경로의 핵심으로 작용하는 효소인, pyruvate decarboxylase(pdc) 유전자의 불활성 실험을 통해, PDC 활성이 50% 감소된 PDC 활성 변형균주가 분리되었다. 이러한 균주들의 에탄올 탄소대사 흐름이 고부가가치 화합물인 피루브산, 숙신산 및 젖산 등으로 전환되는지를 발효 실험을 통해 평가하였다. 하지만 pdc의 발현을 중지시키기 위해 cat-삽입형-pdc와 pdc-결손형 아형 유전자를 전기천공법을 이용해 야생형 균주 ZM4의 염색체에 이식하기 위한 다수의 시도에도 불구하고, 이러한 방법을 통해 분리된 균주들은 대부분 부분적 유전자 불활성 특성을 보였으며, PDC 활성이 완전히 손실된 삭제 돌연변이 균주를 획득할 수는 없었다. PDC활성이 변형된 돌연변이 균주의 발효 실험에서, 야생형 균주와 비교 시 감소된 PDC 효소 활성의 변화로 인해 기질 흡수율과 에탄올 생산율이 감소되어 피루브산 생산이 약 2.5 g l-1 정도로 증가함을 확인하였으나, 젖산과 숙신산의 생산에 현저한 농도 변화를 보이지 못했다. 이러한 결과는 Z. mobilis의 산화환원 에너지가 PDC 효소 활성에 의한 에탄올 생산 경로에 전적으로 의존하여 발생한다는 것을 암시하였다. 상기 결과를 토대로 pdc 유전자의 완전한 불활성 유도와 산화환원 에너지의 균형은, 젖산 생산을 위한 lactate dehydrogenase, 숙신산 생산을 위한 pyruvate dehydrogenase와 malic enzyme과 같은 효소의 활성 증가를 통해, 세포내 NAD와 NADH 농도의 산화환원 균형이 이루어져야 발생할 수 있음을 시사하였다.

Keywords

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