毛果杨羧酸酯酶基因家族的功能研究
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北京林业大学生物科学与技术学院

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国家自然科学基金(No.31570664)


Functional Analysis of Carboxylesterase Gene Family from Populus trichocarpa
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    摘要:

    羧酸酯酶(CXE)是一类具有α/β折叠结构域的水解酶类,在植物生长发育和胁迫响应中发挥着重要作用。本研究从毛果杨基因组中鉴定出52个CXE基因。毛果杨52个CXE基因分为3个类型,每一类型分别有39、4和9个CXE基因。毛果杨基因组中有7个CXE基因簇,分布于6条染色体上,包含有28个CXE基因,其余24个CXE基因分散分布在16个染色体上。毛果杨8个CXE基因在木质部、韧皮部、叶、小枝和根五个组织部位均未检测到表达,其余44个CXE基因在五个组织部位呈现差异性表达。在大肠杆菌中表达并纯化了5个毛果杨CXE蛋白(PtCXE4、5、6、41和43),并测定了5个蛋白对在植物体内发现的24个天然底物的催化能力,发现PtCXE5可以催化天然底物的个数最多(11个),酶学性质分析表明5个蛋白都可以催化4-MU acetate和4-MU butyrate水解,却都不能催化4-MU laurate和4-MU palmitate水解,其随着底物碳链长度的增加,蛋白的催化活性也随之下降。酶活性的pH依赖性分析发现,在pH6.0-9.0范围内随着pH的升高5个蛋白的催化活性均呈现升高的趋势。本研究结果为深入揭示林木CXE基因家族的功能奠定基础。

    Abstract:

    Carboxylesterase (CXE) is widely distributed in plants, and can hydrolyze esters of short-chain fatty acids. CXE belong to α/β hydrolase fold superfamily of proteins, and play key roles in plant growth and development and responses to various stresses. In this study, 52 CXE genes were identified from the Populus trichocarpa genome. These 52 CXE genes were divided into three types, containing 39, 4 and 9 CXE genes, respectively. There are seven CXE clusters on Populus six chromosomes. These seven clusters included 28 CXE genes, and the remaining 24 CXE genes were scattered on Populus 16 chromosomes. Among all the 52 CXEs, expression of eight CXE genes were not detectable in the xylem, phloem, leaf, shooting and root of Populus trichocarpa, while the other 44 CXE genes were differentially expressed in the five tissues. Five CXE proteins (PtCXE4, 5, 6, 41, and 43) were expressed in E. coli, and purified to detect activity. PtCXE5 was found to catalyze the largest number of natural substrates. All the five purified proteins can catalyze the hydrolysis of 4-MU acetate and 4-MU butyrate, but none of them can catalyze the hydrolysis of 4-MU laurate and 4-MU palmitate. The CXE showed lower catalytic activity towards substrates with longer carbon chains.The pH-dependent analysis of enzyme activity showed that the catalytic activity of the five proteins increased with the increase of pH in the range of pH 6.0-9.0. This study provides new insights into the functions of carboxylesterase gene family.

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  • 收稿日期:2020-01-02
  • 最后修改日期:2020-03-24
  • 录用日期:2020-04-03
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