Cloning and functional analysis of a fructosyltransferase cDNA for synthesis of highly polymerized levans in timothy (Phleum pratense L.).
Cloning and functional analysis of a fructosyltransferase cDNA for synthesis of highly polymerized levans in timothy (Phleum pratense L.).
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DOI：
10.1093/jxb/ern337
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发表时间：
2009
影响因子：
6.9
通讯作者：
Yoshida M
中科院分区：
文献类型：
--
作者：
Tamura K;Kawakami A;Sanada Y;Tase K;Komatsu T;Yoshida M
植物果聚糖的结构和聚合度的变化可以解释为具有不同性质的果糖基转移酶（FT）的不同组合的结果。虽然FT基因已经在一系列植物物种中被分离，但蔗糖：果聚糖6-果糖基转移酶（6-SFT）cDNA仅在少数物种如小麦中具有功能特征。从温带牧草梯牧草中鉴定出一个具有6-SFT活性的FT cDNA。从冷驯化的梯牧草中分离出FT同源物PpFT 1的cDNA。在毕赤酵母中表达的重组PpFT 1蛋白显示出6-SFT/蔗糖：蔗糖1-果糖基转移酶（1-SST）活性，并从蔗糖产生线性β（2，6）-连接的果聚糖，其DP比小麦6-SFT（Wft 1）体外形成的禾本科聚糖中存在的DP高。PpFT 1和Wft 1表现出明显不同的受体底物特异性：PpFT 1对6-酮三糖具有高亲和力以产生果聚糖，对1-酮三糖具有低亲和力，而Wft 1优先使用1-酮三糖作为受体。PpFT 1重组酶对底物蔗糖的亲和力低于Wft 1重组酶。它也被证实，梯牧草幼苗在高蔗糖和寒冷条件下的果聚糖积累过程中，PpFT 1转录水平升高。我们的研究结果表明，PpFT 1是一种新的cDNA，具有独特的酶性质，不同于以前克隆的植物6-SFTs，并参与提摩太高度聚合的果聚糖的合成。
Variation in the structures of plant fructans and their degree of polymerization (DP) can be explained as the result of diverse combinations of fructosyltransferases (FTs) with different properties. Although FT genes have been isolated in a range of plant species, sucrose:fructan 6-fructosyltransferase (6-SFT) cDNAs have only been functionally characterized in a few species such as wheat. A novel FT cDNA possessing 6-SFT activity has been identified and characterized from the temperate forage grass, timothy (Phleum pratense L.). The cDNA of an FT homolog, PpFT1, was isolated from cold-acclimated timothy. A recombinant PpFT1 protein expressed in Pichia pastoris showed 6-SFT/sucrose:sucrose 1-fructosyltransferase (1-SST) activity and produced linear β(2,6)-linked levans from sucrose with higher DPs than present in graminans formed in vitro by wheat 6-SFT (Wft1). PpFT1 and Wft1 showed remarkably different acceptor substrate specificities: PpFT1 had high affinity for 6-kestotriose to produce levans and low affinity for 1-kestotriose, whereas Wft1 preferentially used 1-kestotriose as an acceptor. The affinity of the PpFT1 recombinant enzyme for sucrose as a substrate was lower than that of the Wft1 recombinant enzyme. It is also confirmed that timothy seedlings had elevated levels of PpFT1 transcripts during the accumulation of fructans under high sucrose and cold conditions. Our results suggest that PpFT1 is a novel cDNA with unique enzymatic properties that differ from those of previously cloned plant 6-SFTs, and is involved in the synthesis of highly polymerized levans in timothy.
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影响因子：
4.3
作者：
CHATTERTON, NJ;HARRISON, PA;ASAY, KH
通讯作者：
ASAY, KH
影响因子：
3.4
作者：
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通讯作者：
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影响因子：
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作者：
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通讯作者：
HUMPHREYS, MO
影响因子：
4.3
作者：
Kawakami, A;Yoshida, M
通讯作者：
Yoshida, M
影响因子：
4.3
作者：
CHATTERTON, NJ;HARRISON, PA;BENNETT, JH
通讯作者：
BENNETT, JH