Starch Branching Enzyme cDNA from Solanum tuberosum
Starch Branching Enzyme cDNA from Solanum tuberosum
复制标题
马铃薯淀粉分支酶 cDNA
DOI：
10.1104/pp.102.3.1053
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发表时间：
1993
期刊：
影响因子：
4.3
通讯作者：
J. Kreiberg
中科院分区：
文献类型：
--
作者：
P. Poulsen;J. Kreiberg
淀粉是高等植物中主要储存的碳水化合物Ta&1。来自p a t o 的SBE c-NA 的特征，由两种多糖组成，支链淀粉（约75%）和直链淀粉（约25%）。支链淀粉是高度支化的α-1,4葡聚糖，含有α1,6分支点，而直链淀粉由长的线性α-1,4葡聚糖组成，其中一些α-1,6分支点很少。淀粉在叶绿体和淀粉体中通过三种酶活性的作用合成：ADP-Glc 焦磷酸化酶 (EC 2.7.7.27)、淀粉合酶（EC 来源：2.4.1.21）和 SBE (EC 2.4.1.18)（Preiss 综述，1991）。 ADP-Glc焦磷酸化酶催化ADP-Glc的形成，ADP-Glc是淀粉合酶合成直链淀粉的Glc供体，而SBE通过a-1,6分支点将a-1,4葡聚糖的直链添加到直链淀粉来催化直链淀粉转化为支链淀粉。在这里，我们从马铃薯（Solanum tuberosum）中分离出分支酶 cDNA，这是在酶和基因水平上进一步详细研究淀粉生物合成机制的重要先决条件。使用部分马铃薯 SBE cDNA 克隆从马铃薯芽 cDNA 文库中分离出编码 SBE 的全长 cDNA 克隆，该部分马铃薯 SBE cDNA 克隆最初是用编码豌豆 SBE 的异源 cDNA 分离的（表 I）。 3114 bp DNA 序列揭示了一个 ORF，其以坐标 121 bp 处的 ATG 起始密码子开始，以坐标 2704 bp 处的 TGA 密码子结束。 ORF 包含 861 个氨基酸，与玉米和水稻的 SBE I 具有显着相似性（Baba 等，1991；Nakamura 和 Yamanouchi，1992），表明 3.1 kb cDNA 编码马铃薯 SBE。 ORF 编码的蛋白质的计算 M 为 99,083，而据报道，从马铃薯块茎中纯化的分支酶的 M 值在 79,000 至 103,000 范围内（Vos-Scheperkeuter 等，1989；Blennow 和 Johansson，199 1）。对这些差异的理解必须等待对纯化酶的进一步分析。 SBE 已位于造粉体基质中（Kram 等人，1993），这表明推导的 SBE 氨基末端序列包含将 SBE 靶向造粉体的转运肽。根据这一预测，SBE 氨基末端与叶绿体转运肽具有一些共同特征（Gavel 和 von Heine，1990），即高含量的 Ser 和 Thr 残基以及中心带正电荷的结构域。此外，来自马铃薯颗粒结合的淀粉体转运肽的亲水性特征
Starch the main storage carbohydrate in higher plants Ta& 1. Characterjstjcs of the SBE c-NA from p a t o and is composed of the two polysaccharides, amylopectin (approximately 75%) and amylose (approximately 25%). Amylopectin is a highly branched a-1,4 glucan containing a1,6 branch points, whereas amylose is composed of long, linear a-1,4 glucans, some of which have very few a-1,6 branch points. Starch is synthesized in chloroplasts and amyloplasts by the action of three enzymic activities: ADP-Glc pyrophosphorylase (EC 2.7.7.27), starch synthase (EC Source: 2.4.1.21), and SBE (EC 2.4.1.18) (reviewed in Preiss, 1991). ADP-Glc pyrophosphorylase catalyzes the formation of ADP-Glc, which is the Glc donor for the synthesis of amylose carried out by starch synthase, whereas SBE catalyzes the conversion of amylose to amylopectin by adding linear chains of a-1,4 glucans to amylose through a-1,6 branch points. Here we present the isolation of a branching enzyme cDNA from potato (Solanum tuberosum), which is an important prerequisite for further detailed studies of the mechanisms of starch biosynthesis at the enzyme and gene levels. A full-length cDNA clone encoding SBE was isolated from a potato sprout cDNA library using a partial potato SBE cDNA clone that was originally isolated with a heterologous cDNA encoding a pea SBE (Table I). The 3114-bp DNA sequence revealed an ORF that begins with an ATG initiation codon at coordinate 121 bp and ends with a TGA codon at coordinate 2704 bp. The ORF includes 861 amino acids, which has significant similarity to SBE I from maize and rice (Baba et al., 1991; Nakamura and Yamanouchi, 1992), indicating that the 3.1-kb cDNA encodes potato SBE. The protein encoded by the ORF has a calculated M, of 99,083, whereas the purified branching enzyme from potato tubers has been reported to have M, values in the range from 79,000 to 103,000 (Vos-Scheperkeuter et al., 1989; Blennow and Johansson, 199 1). The understanding of these discrepancies must await further analysis of the purified enzyme. SBE has been located in the amyloplast-stroma (Kram et al., 1993), suggesting that the deduced amino-terminal sequence of SBE contains a transit peptide that targets SBE to the amyloplast. In accordance with this prediction, the SBE amino terminus has some features in common with chloroplast transit peptides (Gavel and von Heine, 1990), i.e. a high content of Ser and Thr residues and a central, positively charged domain. Moreover, the hydropathicity profiles of the amyloplast transit peptide from the potato granule-bound