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논문 기본 정보
- 자료유형
- 학술저널
- 저자정보
- 저널정보
- 한국미생물생명공학회 Journal of Microbiology and Biotechnology Journal of Microbiology and Biotechnology 제29권 제4호
- 발행연도
- 2019.1
- 수록면
- 577 - 586 (10page)
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초록· 키워드
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The engineered Aspergillus oryzae has a high NADPH demand for xylose utilization and overproduction of target metabolites. Glucose-6-phosphate dehydrogenase (G6PDH, E.C. 1.1.1.49) is one of two key enzymes in the oxidative part of the pentose phosphate pathway, and is also the main enzyme involved in NADPH regeneration. The open reading frame and cDNA of the putative A. oryzae G6PDH (AoG6PDH) were obtained, followed by heterogeneous expression in Escherichia coli and purification as a his6-tagged protein. The purified protein was characterized to be in possession of G6PDH activity with a molecular mass of 118.0 kDa. The enzyme displayed maximal activity at pH 7.5 and the optimal temperature was 50°C. This enzyme also had a half-life of 33.3 min at 40°C. Kinetics assay showed that AoG6PDH was strictly dependent on NADP+ (Km = 6.3 μM, kcat = 1000.0 s-1, kcat/Km =158.7 s-1·μM-1) as cofactor. The Km and kcat/Km values of glucose-6-phosphate were 109.7 s-1·μM-1 and 9.1 s-1·μM-1 respectively. Initial velocity and product inhibition analyses indicated the catalytic reaction followed a two-substrate, steady-state, ordered BiBi mechanism, where NADP+ was the first substrate bound to the enzyme and NADPH was the second product released from the catalytic complex. The established kinetic model could be applied in further regulation of the pentose phosphate pathway and NADPH regeneration of A. oryzae to improve its xylose utilization and yields of valued metabolites.
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참고문헌 (43)
참고문헌 신청
Olson DG / 2012 / Recent progress in consolidated bioprocessing / Curr. Opin. Biotechnol 23:396~405
Bourdichon F / 2012 / Food fermentations : microorganisms with technological beneficial use / Int. J. Food Microbiol 154:87~97
Szabo OE / 2015 / Ultrasonically assisted single stage and multiple extraction of enzymes produced by Aspergillus oryzae on a lignocellulosic substrate with solid-state fermentation / Biomass Bioenergy 75:161~169
Lin H / 2014 / Engineering Aspergillus oryzae A-4 through the chromosomal insertion of foreign cellulase expression cassette to improve conversion of cellulosic biomass into lipids / PLoS One 9:e108442
El-Ghonemy DH / 2014 / Improvement of Aspergillus oryzae NRRL 3484 by mutagenesis and optimization of culture conditions in solid-state fermentation for the hyper-production of extracellular cellulase / Antonie Van Leeuwenhoek 106:853~864
Bourdichon F / 2012 / Food fermentations : microorganisms with technological beneficial use / Int. J. Food Microbiol 154:87~97
Szabo OE / 2015 / Ultrasonically assisted single stage and multiple extraction of enzymes produced by Aspergillus oryzae on a lignocellulosic substrate with solid-state fermentation / Biomass Bioenergy 75:161~169
Lin H / 2014 / Engineering Aspergillus oryzae A-4 through the chromosomal insertion of foreign cellulase expression cassette to improve conversion of cellulosic biomass into lipids / PLoS One 9:e108442
El-Ghonemy DH / 2014 / Improvement of Aspergillus oryzae NRRL 3484 by mutagenesis and optimization of culture conditions in solid-state fermentation for the hyper-production of extracellular cellulase / Antonie Van Leeuwenhoek 106:853~864
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