基于转录组测序的兔儿伞羽扇豆醇合成途径及关键酶基因研究

为解析兔儿伞羽扇豆醇生物合成途径,探究其关键酶基因,研究运用DNBSEQ测序平台对兔儿伞的叶、茎、根及根茎进行转录组测序,从头组装后获得了191541条Unigenes。KEGG代谢通路分析表明有961条Unigenes涉及兔儿伞羽扇豆醇生物合成途径,其中,395条Unigenes编码羽扇豆醇生物合成途径的17个关键酶。根与其他各组织比较,24条共有差异表达基因涉及羽扇豆醇生物合成途径,编码了法尼基焦磷酸合酶(farnesyl diphosphate synthase,FPPS)、角鲨烯合成酶(squalene synthase,SS)、角鲨烯环氧酶(squalene epoxidase,SE)等关键酶。对关键酶FPPS、SS和SE进行结构分析,发现它们均具有保守的催化结构域和底物结合结构域。研究丰富了兔儿伞植物的功能基因数据库,为进一步研究羽扇豆醇生物合成途径及其关键酶基因的功能和调控机制奠定了基础。

利用AtLUP1超量表达体系在拟南芥中合成羽扇豆醇

羽扇豆醇是一种存在于多种果蔬及中草药中的五环三萜类化合物,具有抗炎症、抗氧化、抗癌等药理作用。羽扇豆醇合酶(Lupeolsynthase,LUP)是羽扇豆醇合成途径中的关键酶。前期研究显示,编码该酶的基因在原核和酵母表达体系中均难以获得有生物活性的蛋白产物。本研究以拟南芥为受体,通过超量表达拟南芥羽扇豆醇合酶1(AtLUP1)基因,分析利用植物细胞生产羽扇豆醇的可行性。使用气相色谱GC检测AtLUP1超量表达拟南芥植株中的羽扇豆醇含量,发现48号植株中羽扇豆醇含量为0.9155mg·g^-1,显著高于野生型。本研究证明超量表达AtLUP1基因可以在拟南芥中促进羽扇豆醇的合成,为羽扇豆醇的工业化生产提供了新的思路。

Current advances in the biotechnological synthesis of betulinic acid:new findings and practical applications

Betulinic acid(BA),a penta-cyclic triterpenoid found as a ubiquitous secondary metabolite throughout the plant kingdom,has aroused tremendous interests due to its different pharmacological properties,which lead to large market demand.However,the content of BA in plant is low for phytoextraction.Although chemical semi-synthesis or biotransformation of BA from betulin with high conversion efficiency is achieved,it still relies on phytoextraction from the bark of medicinal trees.To circumvent this issue,the biotechnological synthesis of BA in engineered yeasts has been developed.In this review,the pharmacological properties of BA are first summarized,including antitumor,anti-HIV,antiprotozoal,anti-inflammatory,apoptosis activator and anti-metabolic syndrome.Then,the traditional phytoextraction,semi-synthesis and biotechnological synthesis of BA are discussed.Particularly,current advances in its biotechnological synthesis and strategies to improve BA production are focused.Moreover,potential strategies for further promotion of BA yield,including the introduction of artificial isopentenol utilization pathway,semi-rational mutagenesis of lupeol synthase and cytochrome P450,and subcellular morphology and compartmentalization,are discussed.

多胺对白桦悬浮细胞生长和三萜积累的影响

目的分析外源多胺对白桦悬浮细胞生长和三萜积累的影响。方法在白桦悬浮细胞的生长末期添加0.1 mmol/L和1.0 mmol/L的腐胺(Put)、精胺(Spm)和亚精胺(Spd),采用比色法和RT-PCR方法分析白桦三萜量及其合成关键酶羽扇豆醇合酶(LUS)基因的表达。结果除1.0 mmol/L Spm处理使白桦悬浮细胞的活力和干质量积累降低外,其他处理均提高了白桦悬浮细胞的活力、干质量积累和三萜产量,且随着处理时间的延长,干质量积累和三萜产量呈增加趋势。其中1.0 mmol/L Put在处理的第2天对细胞干质量和三萜产量的促进效应最大,分别比对照增加了38.89%和116.35%。LUS基因的RT-PCR检测结果进一步证实了多胺对白桦三萜积累的促进作用。结论 Put可有效促进白桦悬浮细胞生长和三萜积累。