A Simple Preparation Method for Phytochromobilin.
A Simple Preparation Method for Phytochromobilin.
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一种植物色素胆素的简便制备方法
DOI：
10.1111/php.12710
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发表时间：
2017
影响因子：
3.3
通讯作者：
Zhao Kai-Hong
中科院分区：
文献类型：
--
作者：
Lu Lu;Zhao Bao-Qing;Miao Dan;Ding Wen-Long;Zhou Ming;Scheer Hugo;Zhao Kai-Hong
植物光敏色素的发色团植物光敏色素移动素（P Φ B）在植物中的浓度很低，很难分离。因此，在植物光敏色素研究中，它经常被蓝藻中丰富的藻蓝胆素所取代。P Φ B也是一个有吸引力的发色团，用于发射远红光的色蛋白。在这项工作中，我们设计并优化了一种简单的方法来有效地分离有用数量的P Φ B：发色团在大肠杆菌中产生，并瞬时结合到ApcE 2的一个定制的发色团结合结构域，ApcE 2是一个核心膜接头的载脂蛋白，随后可以从其中释放。该方法的简易性和有效性不仅取决于在来自集球藻属的ApcE 2构建体的存在下P Φ B的生物合成的增强。PCC7335，但也对色素与其载脂蛋白的非共价结合。分离的P Φ B被成功地整合到光敏色素相关的组装体中，此外，非共价结合的P Φ B可以直接从ApcE2构建体转移到光敏色素、蓝细菌色素和藻胆蛋白的载脂蛋白中，而不会损失相关的生物活性。
Phytochromobilin (PΦB), the chromophore of plant phytochromes, is difficult to isolate because phytochromes occur at very low concentrations in plants. It is, therefore, frequently replaced in plant phytochrome studies by phycocyanobilin, which is abundant in cyanobacteria. PΦB is also an attractive chromophore for far‐red emitting chromoproteins. In this work, we design and optimize a simple method to efficiently isolate useful quantities of PΦB: The chromophore is generated inEscherichia coliand transiently bound to a tailored chromophore‐binding domain of ApcE2, the apo‐protein of a core‐membrane linker, from which it can subsequently be released. The ease and effectiveness of this method hinges not only on the enhanced biosynthesis of PΦB in the presence of the ApcE2 construct fromSynechococcussp. PCC7335, but also on the noncovalent binding of the pigment to its apo‐protein. The isolated PΦB was successfully incorporated into phytochrome‐related assemblies, and furthermore, the noncovalently bound PΦB could be transferred directly from the ApcE2 construct to the apo‐proteins of phytochromes, cyanobacteriochromes and phycobiliproteins, without loss of relevant biological activity.