Vachiranuvathin Phuwanet, Tharasirivat Vetaka, Hemnusornnanon Thitaporn, Jantaro Saowarath
Laboratory of Cyanobacterial Biotechnology, Department of Biochemistry, Faculty of Science, Chulalongkorn University, Bangkok, 10330, Thailand.
Appl Biochem Biotechnol. 2022 Dec;194(12):5748-5766. doi: 10.1007/s12010-022-04061-w. Epub 2022 Jul 11.
To overcome the limited resistance to alcohol stress, genetically engineered Synechocystis sp. PCC 6803 strains with overexpressions of genes related with the ROS detoxification system (sodB and gpx2, which encode superoxide dismutase and glutathione peroxidase, respectively) were developed. Three engineered strains including a sodB-overexpressing strain (OE + S), a gpx2-overexpressing strain (OE + G), and a sodB/gpx2-overexpressing strain (OE + SG) grew similarly as wild-type control under normal condition. When compared to wild-type control, OE + S and OE + SG strains grew faster for 4 days under 2.0% (v/v) ethanol and 0.3% (v/v) n-butanol conditions, as well as having higher chlorophyll a levels. On the other hand, the prominent growth recovery of OE + G and OE + SG was noted within 4 days in normal BG medium after treating cells with high alcohol stresses for 1 h, in particular 15% ethanol and 2.5% n-butanol. Under 4 days of recovery from butanol stress, specific levels of intracellular pigments including chlorophyll a and carotenoids were dramatically increased in all modified strains. The overexpression of antioxidant genes then revealed a significant improvement of alcohol tolerance in Synechocystis sp. PCC 6803.
为了克服对酒精胁迫的有限抗性，构建了基因工程改造的集胞藻6803菌株，这些菌株过表达了与活性氧解毒系统相关的基因（sodB和gpx2，分别编码超氧化物歧化酶和谷胱甘肽过氧化物酶）。包括sodB过表达菌株（OE+S）、gpx2过表达菌株（OE+G）和sodB/gpx2过表达菌株（OE+SG）在内的三种工程菌株在正常条件下与野生型对照生长相似。与野生型对照相比，OE+S和OE+SG菌株在2.0%（v/v）乙醇和0.3%（v/v）正丁醇条件下4天内生长更快，叶绿素a水平也更高。另一方面，在用高酒精胁迫处理细胞1小时后，特别是15%乙醇和2.5%正丁醇，在正常BG培养基中，OE+G和OE+SG在4天内显著恢复生长。在从丁醇胁迫恢复4天后，所有改良菌株中包括叶绿素a和类胡萝卜素在内的细胞内色素的特定水平显著增加。抗氧化基因的过表达随后显示出集胞藻6803对酒精耐受性的显著提高。
