IDENTIFICATION OF A TOMATO GENE FOR THE ETHYLENE-FORMING ENZYME BY EXPRESSION IN YEAST
IDENTIFICATION OF A TOMATO GENE FOR THE ETHYLENE-FORMING ENZYME BY EXPRESSION IN YEAST
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DOI：
10.1073/pnas.88.16.7434
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发表时间：
1991-08-01
影响因子：
11.1
通讯作者：
GRIERSON, D
中科院分区：
文献类型：
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作者：
HAMILTON, AJ;BOUZAYEN, M;GRIERSON, D
乙烯形成酶（EFE）催化植物激素乙烯生物合成的最后一步，从未被纯化过，也没有可用的分子探针。最近，通过在转基因植物中表达的反义基因抑制乙烯合成，已经鉴定了番茄EFE的推定cDNA克隆（pTOM 13）。已通过在酿酒酵母中表达pTOM 13基因对其功能进行了直接测试。在cDNA的5 ′区发现克隆伪迹后，通过将基因组克隆的5 ′端与cDNA的3 ′端融合产生校正的cDNA（pRC 13），并在S.啤酒。转化的酵母培养物将1-氨基环丙烷-1-羧酸（ACC）转化为乙烯，而对照细胞则没有。这种EFE活性显示出与植物组织中发现的EFE相似的特征：它将ACC类似物1-氨基-2-乙基环丙烷-1-羧酸的反式异构体优先于顺式异构体转化为1-丁烯，并且它被钴离子和1，10-菲咯啉强烈抑制。此外，从酵母EFE活性的效应子的活性获得的信息支持EFE是羟化酶组之一的假设。
The ethylene-forming enzyme (EFE), which catalyzes the last step in the biosynthesis of the plant hormone ethylene, has never been purified and no molecular probes are available. Recently, a putative cDNA clone for tomato EFE (pTOM13) has been identified by inhibiting ethylene synthesis with an antisense gene expressed in transgenic plants. A direct test of its function has been made by expression of a pTOM13 gene in Saccharomyces cerevisiae. After cloning artefacts were discovered in the 5' region of the cDNA, a corrected cDNA (pRC13) was created by the fusion of the 5' end of a genomic clone to the 3' end of the cDNA and expressed in S. cerevisiae. Cultures of transformed yeast converted 1-aminocyclopropane-1-carboxylic acid (ACC) to ethylene, whereas control cells did not. This EFE activity displays similar characteristics to EFE found in plant tissue: it converts the trans isomer of the ACC analogue 1-amino-2-ethylcyclopropane-1-carboxylic acid to 1-butene in preference to the cis isomer, and it is strongly inhibited by cobaltous ions and 1,10-phenanthroline. Furthermore, information gained from the activity of effectors on yeast EFE activity supports the hypothesis that EFE is one of a group of hydroxylase enzymes.