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Saccharomyces cerevisiae 에서 Aspergillus oryzae 유래의 exo-β-1,3-glucanase (laminarinase)의 생산 최적화

원문정보

Optimization for Production of Exo-β-1,3-glucanase (Laminarinase) from Aspergillus oryzae in Saccharomyces cerevisiae

김민정, 남수완, Koichi Tamano, Masayuki Machida, 김성구, 김연희

피인용수 : 0(자료제공 : 네이버학술정보)

초록

영어

In this study, a EXGA gene code for exo-β-1,3- glucanase from Aspergillus oryzae was overexpressed and secretory produced in Saccharomyces cerevisiae. To overexpress the β-1,3-glucanase, pGInu-exgA and pAInu-exgA plasmids having GAL10 and ADH1 promoter, respectively, and exoinulinase signal sequence (Inu s.s) were constructed and introduced in S. cerevisiae SEY2102 and 2805. The recombinant β-1,3- glucanase was successfully expressed and secreted into the medium and the β-1,3-glucanase activity in 2102/pGInu-exgA and 2102/pAInu-exgA strain were 5.01 unit/mL and 4.09 unit/mL, respectively. In the 2805/pGInu-exgA and 2805/pAInu-exgA strain, the β-1,3-glucanase activity showed 3.23 unit/mL and 3.22 unit/mL, respectively. Secretory efficiency in each strain reached 95% to 98%. Subsequently, the recombinant β-1,3-glucanase was used for ethanol production. Ethanol productivity in 2102/pAInu-exgA strain was 0.83 g/L when pre-treated Laminaria japonica which has initial reducing sugar of 1.4 g/L was used as substrate. It is assumed that the polysaccharides of Laminaria japonica was effectively saccharified by recombinant β-1,3-glucanase, resulting in increase of ethanol productivity. These results suggested that recombinant β-1,3- glucanase was efficiently overexpressed and secreted in S. cerevisiae SEY2102 as host strain by using ADH1 promoter-Inu s.s system.

목차

Abstract
 1. 서론
 2. 재료 및 방법
  2.1. 사용균주 및 plasmids
  2.2. 재조합 plasmid 구축 및 형질전환
  2.3. 배지 조성 및 배양조건
  2.4. 균체 농도 및 Plasmid 안정성
  2.5. MUG plate assay
  2.6. 균체분획 및 β-1,3-glucanase 활성 측정과 분비효율
  2.7. SDS-PAGE에 의한 단백질 확인
  2.8. 에탄올 생산성 측정
 3. 결과 및 고찰
  3.1. β-1,3-glucanase 발현 재조합 plasmid의 구축
  3.2. 효모 형질전환체 선별 및 MUG assay
  3.3. β-1,3-glucanase 과발현 및 분비생산
  3.4. β-1,3-glucanase 단백질 확인 및 에탄올 생산성
 4. 결론
 감사
 References

저자정보

  • 김민정 Min-Jung Kim. 동의대학교 바이오물질제어학과
  • 남수완 Soo-Wan Nam. 동의대학교 바이오물질제어학과, 동의대학교 생명공학과
  • Koichi Tamano 일본 산업기술종합연구소 (AIST)
  • Masayuki Machida 일본 산업기술종합연구소 (AIST)
  • 김성구 Sung-Koo Kim. 부경대학교 생물공학과
  • 김연희 Yeon-Hee Kim. 동의대학교 바이오물질제어학과, 동의대학교 생명공학과

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