Department of Biotechnology, Catholic University of Korea, Bucheon, Gyeonggi, Korea.
Lett Appl Microbiol. 2011 May;52(5):433-40. doi: 10.1111/j.1472-765X.2011.03013.x. Epub 2011 Feb 15.
To understand the intracellular reducing power metabolism, growth and intracellular NAD(P)H concentrations of a phosphoglucose isomerase (pgi)-disrupted Escherichia coli (KS002) were investigated with the expressions of redox enzymes.
The isogenic pgi-mutation enabled E. coli to harbour two times both the intracellular NADPH and NADH at half the growth rate. The wild-type expressing NAD-dependent malic enzyme (maeA) was incapable of sufficient growth (<0·02 h(-1)), and the growth retardations were distinctively recovered when NADP-dependent glyceraldehyde-3-phosphate dehydrogenase (gapB) from Bacillus subtilis was coexpressed. The KS002 expressing maeA harboured the highest intracellular reducing powers (NADPH of 3·9 and NADH of 5·2 μmol g DCW(-1) ) by three times each of those in wild type. The expression of NADP-dependent malic enzyme (maeB) enabled wild-type and KS002 strains to grow without significant alteration.
The alterations of reducing powers and the growth were analysed in the genetic engineered E. coli strains. The potential application of the cells with the high intracellular NAD(P)H level is discussed based on the results.
Metabolic engineering strategy for higher reducing power regeneration is provided.
通过对氧化还原酶的表达，研究磷酸葡萄糖异构酶（pgi）缺失的大肠杆菌（KS002）的细胞内还原力代谢、生长和细胞内 NAD(P)H 浓度。
同基因 pgi 突变使大肠杆菌能够以半生长速率保持两倍的细胞内 NADPH 和 NADH。表达 NAD 依赖性苹果酸酶（maeA）的野生型几乎无法生长（<0·02 h(-1)），当共表达来自枯草芽孢杆菌的 NADP 依赖性甘油醛-3-磷酸脱氢酶（gapB）时，生长迟缓明显恢复。表达 maeA 的 KS002 具有最高的细胞内还原力（NADPH 为 3.9 μmol g DCW(-1)，NADH 为 5.2 μmol g DCW(-1)），分别是野生型的三倍。NADP 依赖性苹果酸酶（maeB）的表达使野生型和 KS002 菌株能够生长，而没有明显的变化。
分析了遗传工程大肠杆菌菌株的还原力变化和生长情况。根据结果讨论了具有高细胞内 NAD(P)H 水平的细胞的潜在应用。
提供了更高还原力再生的代谢工程策略。
