In vitro Biochemical Characterization of All Barley Endosperm Starch Synthases.
In vitro Biochemical Characterization of All Barley Endosperm Starch Synthases.
复制标题
DOI：
10.3389/fpls.2015.01265
复制
发表时间：
2015
影响因子：
5.6
通讯作者：
Palcic MM
中科院分区：
文献类型：
--
作者：
Cuesta-Seijo JA;Nielsen MM;Ruzanski C;Krucewicz K;Beeren SR;Rydhal MG;Yoshimura Y;Striebeck A;Motawia MS;Willats WG;Palcic MM
淀粉是谷物中主要的贮藏多糖，也是人类饮食中主要的热量来源。它由一组酶合成，包括五类淀粉酶（SS）。虽然总体淀粉合成酶（SS）的反应是已知的，五个SS类之间的功能差异知之甚少。我们的大部分知识来自分析突变体植物与改变SS活动，但由此产生的数据往往是难以解释的结果，多效效应，酶之间的竞争，重叠的酶活性和多酶复合物的破坏。在这里，我们提供了一个详细的生化研究的所有五类SS在大麦胚乳的活动。每种酶都是在E.大肠杆菌和性质和模式的作用在体外进行了研究，从其他SS和其他底物修饰活动的隔离。我们的研究结果定义了每个SS类前所未有的详细的行动模式，我们分析了他们的基板选择，温度依赖性和稳定性，基板亲和力和时间丰富的大麦发展。我们的研究结果与一些普遍接受的淀粉生物合成的想法是不一致的，并可能导致在植物中获得的结果重新解释。特别是，他们指出，颗粒结合的SS能够进行性行动，即使在没有淀粉基质，SSI没有伸长率限制，和SSIV，认为是关键的启动淀粉颗粒，麦芽糖而不是多糖作为其优选的底物。
Starch is the main storage polysaccharide in cereals and the major source of calories in the human diet. It is synthesized by a panel of enzymes including five classes of starch synthases (SSs). While the overall starch synthase (SS) reaction is known, the functional differences between the five SS classes are poorly understood. Much of our knowledge comes from analyzing mutant plants with altered SS activities, but the resulting data are often difficult to interpret as a result of pleitropic effects, competition between enzymes, overlaps in enzyme activity and disruption of multi-enzyme complexes. Here we provide a detailed biochemical study of the activity of all five classes of SSs in barley endosperm. Each enzyme was produced recombinantly in E. coli and the properties and modes of action in vitro were studied in isolation from other SSs and other substrate modifying activities. Our results define the mode of action of each SS class in unprecedented detail; we analyze their substrate selection, temperature dependence and stability, substrate affinity and temporal abundance during barley development. Our results are at variance with some generally accepted ideas about starch biosynthesis and might lead to the reinterpretation of results obtained in planta. In particular, they indicate that granule bound SS is capable of processive action even in the absence of a starch matrix, that SSI has no elongation limit, and that SSIV, believed to be critical for the initiation of starch granules, has maltoligosaccharides and not polysaccharides as its preferred substrates.