摘要
香叶基焦磷酸合成酶(GPS)在植物体内催化二甲烯丙基焦磷酸与异戊烯基焦磷酸结合成萜类化合物前香叶基香叶基焦磷酸(GPP)。为深入理解GPS蛋白在类胡萝卜素生物合成过程中的作用,本研究采用高通量测序方法获得绿色杜氏藻GPS基因(Gen Bank序号:KT751181)c DNA全长序列,并研究乙酰水杨酸(ASA)、硫酸铈铵(CAS)和花生四烯酸(AA)3种诱导子对绿色杜氏藻GPS基因表达的影响。测序结果表明,绿色杜氏藻GPS基因c DNA全长2 281 bp,开放阅读框为1 449 bp,编码463个氨基酸。绿色杜氏藻GPS蛋白的分子量为53 714.9,理论等电点(p I)为6.66,该蛋白分子呈亲水性,无信号肽和跨膜结构域。二级结构分析显示,该蛋白分子中螺旋结构占主导地位,占47.7%。系统进化树结果表明,绿色杜氏藻GPS蛋白与莱茵衣藻亲缘关系最相近。诱导表达分析显示,62.5 mg·L-1AA、0.2~0.8 mg·L-1ACS和10 mg·L-1ASA可极显著提高绿色杜氏藻GPS基因表达;同时,类胡萝卜素含量也极显著升高,这一结果表明GPS基因与类胡萝卜素合成有关,GPS基因可能通过控制GPP的去向,间接影响绿色杜氏藻类胡萝卜素合成。本研究结果为揭示藻类类胡萝卜素合成机理及利用代谢工程策略提高类胡萝卜素含量奠定了理论基础。
Geranyl pyrophosphate synthase catalyzes dimethyl allyl pyrophosphate reacting with iso-amyl alkenyl pyrophosphate to synthesize Geranyl pyrophosphate, a kind of terpenoid precursors in plant. In order to understand more about the role of GPS involved in the carotenoid biosynthesis at the molecular level. In this study, the eDNA full length of GPS gene （GenBank accession number： KT751181 ） in Dunaliella viridis was obtained. Aacetylsalicylic acid （ASA） , ammonium cerous sulfate （ACS） and arachidonic acid （AA） were used to induce the expression of GPS gene in D. viridis. The GPS gene of D. viridis had a full length of 2281 bp, containing an open reading frame of 1449 bp, encoding 463 amino acids. The relative molecular mass of GPS protein was 53714.9, the theoretical isoelectric point （PI） was 6.66. Bioinformatic prediction of GPS protein revealed that the protein was hydrophilie, and had neither signal peptide nor transmembrane domain. The secondary structure analysis illustrated that approximately 47.7% of GPS was helix structure. Phylogenetic tree results showed that the protein sequence of GPS of D. viridis was similar to that of Chlamydomonas reinhardtii. The induced expression analysis implied that transcription levels of GPS gene in D. viridis significantly increased when treated by 62.5 mg·L-1AA, 0.2 to 0. 8 mg·L-1ACS and 10 mg·L-1ACS. Meanwhile, the content of carotenoids was also increased significantly. Therefore, GPS gene was related to the biosynthesis of carotenoid. GPS gene may indirectly affect the carotenoid content, by controlling GPP synthesis. This study will provide theoretical basis on molecular mechanism of carotenoid biosynthesis study, and improving content of carotenoid though metabolic engineering strategies in algae.
出处
《核农学报》
CAS
CSCD
北大核心
2017年第2期248-254,共7页
Journal of Nuclear Agricultural Sciences
基金
浙江省科技厅重点科技创新团队项目(2012R10029-07
2010R50029)
宁波科技攻关项目(2013C10018)
