Korea Institute of Ocean Science and Technology, Ansan, 426-744, South Korea.
Appl Microbiol Biotechnol. 2014 Jan;98(2):979-86. doi: 10.1007/s00253-013-5448-y. Epub 2013 Dec 13.
To overproduce biotechnologically valuable products, the expression level of target genes has been modulated by using strong promoters. In a hyperthermophilic archaeon Thermococcus onnurineus NA1, two promoters, P(TN0413) and P(TN0157), which drive expression of the genes encoding the S-layer protein and glutamate dehydrogenase were inserted in front of a gene cluster encoding a carbon monoxide dehydrogenase, a hydrogenase and a Na⁺/H⁺ antiporter. Two promoters exhibited strong activity by increasing the transcription and translation levels of the gene cluster in the mutant strains by 2.5- to 49-folds and 1.4- to 3.3-folds, respectively, than the native promoter in the wild-type strain. While KS0413 with P(TN0413) promoter exhibited 2.7 to 4.7 times higher transcript level than KS0157 with P(TN0157) promoter, the levels of proteins were a little different between them. The biomass concentrations and H₂ production rates of two mutants were 2- to 3-fold higher than those of the wild-type strain in a bioreactor where CO was supplied at a flow rate of 120 ml min⁻¹. Two mutants showed differential response to the higher CO flow rate, 240 ml min⁻¹, in terms of growth pattern and product formation, indicating two promoters were regulated by culture conditions. The results demonstrate that not only promoter strength but also product-forming conditions should be considered in promoter engineering.
为了大量生产具有生物技术价值的产品，人们通过使用强启动子来调节目标基因的表达水平。在一种嗜热古菌 Thermococcus onnurineus NA1 中，两个启动子 P(TN0413) 和 P(TN0157) 被插入到编码表面层蛋白和谷氨酸脱氢酶的基因之前，分别驱动一氧化碳脱氢酶、氢化酶和 Na⁺/H⁺反向转运体的基因簇的表达。与野生型菌株中的天然启动子相比，这两个启动子分别将基因簇的转录和翻译水平提高了 2.5-49 倍和 1.4-3.3 倍，从而使突变株中的基因表达水平显著增强。虽然带有 P(TN0413) 启动子的 KS0413 的转录水平比带有 P(TN0157) 启动子的 KS0157 高 2.7-4.7 倍，但两者的蛋白水平略有不同。在以 120 ml min⁻¹ 的流速供应 CO 的生物反应器中，两个突变体的生物量浓度和 H₂ 生成速率比野生型菌株高 2-3 倍。两个突变体对更高的 CO 流速（240 ml min⁻¹）表现出不同的生长模式和产物形成的响应，表明两个启动子受到培养条件的调节。这些结果表明，在启动子工程中，不仅要考虑启动子的强度，还要考虑产物形成的条件。
