Functional immobilization of plant receptor-like kinase onto microbeads towards receptor array construction and receptor-based ligand fishing.
Functional immobilization of plant receptor-like kinase onto microbeads towards receptor array construction and receptor-based ligand fishing.
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DOI：
10.1111/j.1365-313x.2007.03204.x
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发表时间：
2007-07
期刊：
影响因子：
--
通讯作者：
Hidefumi Shinohara;Y. Matsubayashi
中科院分区：
文献类型：
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作者：
Hidefumi Shinohara;Y. Matsubayashi
尽管在植物中存在大量的受体样激酶（RLKs），但很少有其特异性配体是已知的。配体钓鱼是最具挑战性的后基因组技术之一。在这里，我们报告了一种策略，通过共价键在微珠上的植物RLKs的功能性固定。由于固定化的RLKs的高密度，配体-受体相互作用可以在共聚焦激光扫描显微镜下使用荧光标记的配体以高灵敏度可视化。此外，使用基于受体的亲和层析系统与RLK微珠，受体的配体直接从复杂的天然样品中以高纯度回收。我们的RLK微珠和基于受体的配体捕捞方法是一种可行的替代传统的正向遗传学和基于生物测定的生化纯化，用于鉴定植物中的新型配体-受体对。
Despite the large number of receptor-like kinases (RLKs) in plants, few of their specific ligands are known. Ligand fishing is one of the most challenging post-genomic technologies. Here, we report a strategy for functional immobilization of plant RLKs on microbeads via covalent linkage. Because of the high density of immobilized RLKs, ligand-receptor interaction can be visualized at high sensitivity using fluorescent-labeled ligands under the confocal laser scanning microscope. Moreover, using a receptor-based affinity chromatography system with RLK microbeads, the ligand of the receptor was directly retrieved at high purity from complex natural samples. Our RLK microbeads and receptor-based ligand fishing approach is a feasible alternative to conventional forward genetics and bioassay-based biochemical purification for identification of novel ligand-receptor pairs in plants.