佐夫色绿藻高产虾青素突变体的筛选及虾青素合成机理研究  

SCREENING OF CHROMOCHLORIS ZOFINGIENSIS HYPER ASTAXANTHINPRODUCING MUTANTS AND THE MECHANISMS UNDERLYING ASTAXANTHIN SYNTHESIS

作　　者：肖玲 韩丹翔[1] XIAO Ling;HAN Dan-Xiang(Center for Microalgal Biotechnology and Biofuels,Institute of Hydrobiology,Chinese Academy of Sciences,Wuhan 430072,China;University of Chinese Academy of Sciences,Beijing 100049,China)

机构地区：[1]中国科学院水生生物研究所藻类生物技术和生物能源研发中心,武汉430072 [2]中国科学院大学,北京100049

出　　处：《水生生物学报》2023年第1期92-103,共12页Acta Hydrobiologica Sinica

基　　金：国家重点研发计划合成生物学专项(2018YFA0902500)资助。

摘　　要：为了提高佐夫色绿藻(Chromochloris zofingiensis)细胞内虾青素含量,研究通过甲基磺酸乙酯诱变构建了含有20000个单克隆的突变体库,并筛选出一株高产虾青素的突变体12C10。在异养正常培养条件下,当葡萄糖耗完时,12C10虾青素含量比野生型提高74%;缺氮诱导第4天时,虾青素含量比野生型高25%。利用广泛靶向代谢组学分析在正常培养条件下12C10与野生型在代谢物水平上的差异。与野生型相比,12C10中除谷氨酸外的氨基酸及脂肪酸的合成普遍下降,但是谷氨酸的水平显著提高。氨基酸和脂肪酸合成减少为虾青素合成提供更多的碳骨架、NADPH和ATP。谷氨酸的积累可能一方面刺激了磷酸戊糖途径中6-磷酸葡萄糖脱氢酶的活性促进NADPH的产生,另一方面导致氧自由基产生促进虾青素合成。代谢物组分析结果还表明12C10中虾青素合成的增强可能与乙烯合成的增强有关。研究为进一步通过代谢调控提高C.zofingiensis虾青素含量奠定了基础,对指导C.zofingiensis虾青素积累新工艺的开发具有重要意义。Chromochloris zofingiensis can achieve ultra-high cell density and simultaneously accumulate astaxanthin under heterotrophic conditions, however, the relatively low cellular astaxanthin content of C. zofingiensis hinders its commercial application. With the aim to improve the ellular astaxanthin content of C. zofingiensis, this study screened a mutant 12C10 with enhanced astaxanthin content from the mutant library established by using ethylmethylsulfone mutagenesis. The astaxanthin content of 12C10 was 74% higher than that of the wild type under the heterotrophic normal conditions, and 25% higher than that of the wild type at day 4 under the nitrogen deficiency conditions. Additionally, the results of the widely targeted metabolomics analysis revealed the differences in the metabolites level between12C10 and wild type under the normal conditions. Specifically, the contents of fatty acids and amino acids except glutamate generally decreased, but the content of glutamate significantly increased in 12C10 compared to wild type.Reduced synthesis of amino acids and fatty acids in 12C10 can provide more carbon skeleton, NADPH, and ATP for astaxanthin synthesis. The accumulation of glutamate on the one hand may stimulate the activity of glucose 6-phosphate dehydrogenase in pentose-phosphate pathway to produce more NADPH, and cause production of reactive oxygen species to trigger astaxanthin synthesis. The metabolome analysis also showed that the enhanced astaxanthin synthesis in 12C10 may be related to the enhanced ethylene synthesis. This study lays the foundation for improving C. zofingiensis astaxanthin content through metabolic regulation in the future, and is of great significance for guiding the development of the new processes of C. zofingiensis astaxanthin accumulation.

关 键 词：EMS诱变 虾青素突变体 差异代谢物 佐夫色绿藻 

分 类 号：Q949.2[生物学—植物学]