微拟球藻Ⅰ型脂酰辅酶A:二脂酰甘油酰基转移酶编码基因NoDGAT1A的功能分析

Functional verification of an acyl-Co A: diacylglycerol acyltransferase 1-like gene(NoDGAT1A) from oleaginous microalga Nannochloropsis oceanica

从微拟球藻高产油藻株IMET1的cDNA中,克隆出一个I型脂酰辅酶A:二脂酰甘油酰基转移酶的编码基因NoDGAT1A,该基因共编码437个氨基酸,与拟南芥I型DGAT的氨基酸序列相似度为38%,且含有至少9段高疏水区。随后,NoDGAT1A被转化入营养缺陷型酿酒酵母三脂酰甘油合成突变株H1246中进行诱导表达。结果表明,在外加二十碳五烯酸(EPA)时,表达No DGAT1A的H1246能够产生三脂酰甘油(TAG),针对其TAG的分析表明,与酵母DGAT相比,表达NoDGAT1A的H1246产生的TAG中含有更多的EPA,说明NoDGAT1A具有更强的合成含EPA的TAG之能力。此外,16:0(63.82%)和18:0(27.98%)是其TAG中主要的脂肪酸链成分,说明NoDGAT1A合成的TAG侧链仍以长链饱和脂肪酸为主。NoDGAT1A对微拟球藻中TAG的合成具有重要作用,也为以微拟球藻为模式的产油微藻TAG和EPA代谢网络提供参考。

We identified and isolated a cDNA which encodes a novel acyl-CoA： diacylglycerolacyl transferase （DGAT） 1-like protein NoDGAT1A, from the oleaginous microalgaNannochloropsis oceanicaIMET1.NoDGAT1A encodes a 437-amino acid polypeptide that is 38% identical to itsArabidopsis thaliana counterpart and harbors at least nine hydrophobic domains. In aSaccharomyces cerevisiae TAG-deficient mutant H1246, the DGAT activity was restored when expressing NoDGAT1A and supplementing eicosapentaenoic acid （EPA） as an additional sub-strate. Moreover, NoDGAT1A incorporated more EPA into TAGs than yeast DGAT, suggesting a higher produc-tivity of EPA-containing TAG species by NoDGAT1A. Furthermore, 16： 0 （63.82%） and 18： 0 （27.98%） are the dominant fatty acid chains in the TAGs synthesized by NoDGAT1A, suggesting that most of the TAGs produced by NoDGAT1A are made up of long chain saturated fatty acids. The evidence supports a role of NoDGAT1A in TAG synthesis inN. oceanica and has implications in understanding the functional network underlying TAG and EPA synthesis in microalgae.