Takahashi Shigekazu, Abe Eriko, Nakayama Katsumi, Satoh Hiroyuki
a Faculty of Science, Department of Biomolecular Science , Toho University , Funabashi , Japan.
Biosci Biotechnol Biochem. 2015;79(2):205-10. doi: 10.1080/09168451.2014.972326. Epub 2014 Oct 21.
Photoconvertible water-soluble chlorophyll-binding proteins, called Class I WSCPs, have been detected in Chenopodiaceae, Amaranthaceae and Polygonaceae plant species. To date, Chenopodium album WSCP (CaWSCP) is the only cloned gene encoding a Class I WSCP. In this study, we identified two cDNAs encoding Chenopodium ficifolium Class I WSCPs, CfWSCP1, and CfWSCP2. Sequence analyses revealed that the open reading frames of CfWSCP1 and CfWSCP2 were 585 and 588 bp, respectively. Furthermore, both CfWSCPs contain cystein2 and cystein30, which are essential for the chlorophyll-binding ability of CaWSCP. Recombinant CfWSCP1 and CfWSCP2, expressed in Escherichia coli as hexa-histidine fusion proteins (CfWSCP1-His and CfWSCP2-His), formed inclusion bodies; however, we were able to solubilize these using a buffer containing 8 M urea and then refold them by dialysis. The refolded CfWSCP1-His and CfWSCP2-His could bind chlorophylls and exhibited photoconvertibility, confirming that the cloned CfWSCPs are further examples of Class I WSCPs.
在藜科、苋科和蓼科植物物种中已检测到可光转换的水溶性叶绿素结合蛋白，即I类水溶性叶绿素结合蛋白（WSCPs）。迄今为止，藜菜WSCP（CaWSCP）是唯一克隆的编码I类WSCP的基因。在本研究中，我们鉴定出两个编码藜叶藜I类WSCP的cDNA，即CfWSCP1和CfWSCP2。序列分析表明，CfWSCP1和CfWSCP2的开放阅读框分别为585和588 bp。此外，两种CfWSCP均含有半胱氨酸2和半胱氨酸30，这对CaWSCP的叶绿素结合能力至关重要。在大肠杆菌中作为六聚组氨酸融合蛋白（CfWSCP1-His和CfWSCP2-His）表达的重组CfWSCP1和CfWSCP2形成了包涵体；然而，我们能够使用含有8 M尿素的缓冲液将它们溶解，然后通过透析使其复性。复性后的CfWSCP1-His和CfWSCP2-His能够结合叶绿素并表现出光转换能力，证实克隆的CfWSCPs是I类WSCP的更多实例。
