Biosynthesis of fluorescent cyanobacterial allophycocyanin trimer in Escherichia coli

doi:10.1007/s11120-010-9574-4

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	Biosynthesis of fluorescent cyanobacterial allophycocyanin trimer in Escherichia coli
	Liu, Shaofang 1,2; Chen, Yingjie 1,2; Lu, Yandu 2,3; Chen, Huaxin 1; Li, Fuchao 1; Qin, Song 1
	2010-08-01
发表期刊	PHOTOSYNTHESIS RESEARCH
ISSN	0166-8595
卷号	105 期号:2 页码:135-142
文章类型	Article
摘要	Allophycocyanin (APC), a cyanobacterial photosynthetic phycobiliprotein, functions in energy transfer as a light-harvesting protein. One of the prominent spectroscopic characteristics of APC is a strong red-shift in the absorption and emission maxima when monomers are assembled into a trimer. Previously, holo-APC alpha and beta subunits (holo-ApcA and ApcB) were successfully synthesized in Escherichia coli. In this study, both holo-subunits from Synechocystis sp. PCC 6803 were co-expressed in E. coli, and found to self-assemble into trimers. The recombinant APC trimer was purified by metal affinity and size-exclusion chromatography, and had a native structure identical to native APC, as determined by characteristic spectroscopic measurements, fluorescence quantum yield, tryptic digestion analysis, and molecular weight measurements. Combined with results from a study in which only the monomer was formed, our results indicate that bilin synthesis and the subsequent attachment to apo-subunits are important for the successful assembly of APC trimers. This is the first study to report on the assembly of recombinant ApcA and ApcB into a trimer with native structure. Our study provides a promising method for producing better fluorescent tags, as well as a method to facilitate the genetic analysis of APC trimer assembly and biological function.; Allophycocyanin (APC), a cyanobacterial photosynthetic phycobiliprotein, functions in energy transfer as a light-harvesting protein. One of the prominent spectroscopic characteristics of APC is a strong red-shift in the absorption and emission maxima when monomers are assembled into a trimer. Previously, holo-APC alpha and beta subunits (holo-ApcA and ApcB) were successfully synthesized in Escherichia coli. In this study, both holo-subunits from Synechocystis sp. PCC 6803 were co-expressed in E. coli, and found to self-assemble into trimers. The recombinant APC trimer was purified by metal affinity and size-exclusion chromatography, and had a native structure identical to native APC, as determined by characteristic spectroscopic measurements, fluorescence quantum yield, tryptic digestion analysis, and molecular weight measurements. Combined with results from a study in which only the monomer was formed, our results indicate that bilin synthesis and the subsequent attachment to apo-subunits are important for the successful assembly of APC trimers. This is the first study to report on the assembly of recombinant ApcA and ApcB into a trimer with native structure. Our study provides a promising method for producing better fluorescent tags, as well as a method to facilitate the genetic analysis of APC trimer assembly and biological function.
关键词	Allophycocyanin Trimer Assembly Recombinant Protein Allophycocyanin Monomer Fluorescence Escherichia Coli
学科领域	Plant Sciences
DOI	10.1007/s11120-010-9574-4
URL	查看原文
收录类别	SCI
语种	英语
WOS记录号	WOS:000280921200006
引用统计	被引频次：24[WOS] [WOS记录] [WOS相关记录]
文献类型	期刊论文
条目标识符	http://ir.qdio.ac.cn/handle/337002/5792
专题	实验海洋生物学重点实验室
作者单位	1.Chinese Acad Sci, Key Lab Expt Marine Biol, Inst Oceanol, Qingdao 266071, Peoples R China 2.Chinese Acad Sci, Grad Sch, Beijing 100049, Peoples R China 3.Chinese Acad Sci, Yantai Inst Coastal Zone Res, Yantai 264003, Peoples R China
第一作者单位	中国科学院海洋研究所
推荐引用方式 GB/T 7714	Liu, Shaofang,Chen, Yingjie,Lu, Yandu,et al. Biosynthesis of fluorescent cyanobacterial allophycocyanin trimer in Escherichia coli[J]. PHOTOSYNTHESIS RESEARCH,2010,105(2):135-142.
APA	Liu, Shaofang,Chen, Yingjie,Lu, Yandu,Chen, Huaxin,Li, Fuchao,&Qin, Song.(2010).Biosynthesis of fluorescent cyanobacterial allophycocyanin trimer in Escherichia coli.PHOTOSYNTHESIS RESEARCH,105(2),135-142.
MLA	Liu, Shaofang,et al."Biosynthesis of fluorescent cyanobacterial allophycocyanin trimer in Escherichia coli".PHOTOSYNTHESIS RESEARCH 105.2(2010):135-142.

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