其他题名Structural and functional investigation of allophycocyanin trimer
李文军
; 蒲洋; 牛壮; 薛春岭; 秦松发表期刊科学通报
ISSN0023-074X
2017-06-10
卷号62期号:16页码:1699-1713
关键词recombinant allophycocyanin trimerexciton-coupled pairsabsorption spectrumfluorescence spectrumcircular dichroism spectraconformational stability重组别藻蓝蛋白三聚体激子耦合对吸收光谱荧光光谱圆二色谱构象稳定性
DOIdoi: 10.1360/N972016-01416
产权排序(1) 中国科学院烟台海岸带研究所; (2) 中国科学院大学; (3) 鲁东大学农学院
作者部门海岸带生物学与生物资源保护实验室
英文摘要Nature has been developing and optimizing photosynthesis billions years, including green plants, algae, photosynthetic bacteria and cyanobacteria. Phycobiliproteins system, as the phycobilisomes, in cyanobacteria and red algae is the one of two major light-harvesting systems, after evolution in billions years. Allophycocyanin (APC) trimer is a primary composition of phycobilisomes core for efficient lower-energy photons funneling pathway. In 2009, Liu et al. have firstly assembled the highly soluble self-assembled recombinant APC trimer with N-terminus polyhistidine, which the entire biosynthesis pathway of APC trimer was reconstituted in Escherichia coli strain with the full-length genes of apoprotein a subunit and b subunit from Synechocystis sp. PCC 6803. The recombinant APC trimer was used to be the subject in this research. An optimum protocol with immobilized metal-ion affinity chromatography with chelating transition metal ion (Ni~(2+)) was used to purify the recombinant protein. Then, the second purification was performed to obtain APC trimer with a higher purity (A650/A620=1.346), using density gradient centrifugation. Single molecules of recombinant APC trimers were investigated with atomic force microscopy. Three-dimensional images with resolution at nanoscale indicated that the recombinant allophycocyanin trimers were approximately 1.5 nm in short axis, and 15 nm in long axis, which were in accordance with the sizes of natural allophycocyanin trimer measured by transmission electron microscopy or X-ray crystallography. Moreover, the correct conformation of recombinant allophycocyanin trimer was confirmed by fluorescence spectra (studied energy transfer ability), absorption spectra (studied the light-harvesting ability), and circular dichroism (CD) spectra (studied changes of secondary structure and aggregation state). The CD spectra of recombinant allophycocyanin induced by a wide range of pH were analyzed both at wavelengths in ultraviolet range (190-250 nm) and visible range (450-750 nm). The a-helices were the dominant secondary structure in recombinant allophycocyanin trimer by analysis of CD spectra in ultraviolet range, while the analysis of CD spectra in visible range suggested the existence of exciton-coupled pairs of phycocyanobilins, in which the energy transfer was mainly in the form of exciton splitting. By studying its spectra property variations in response to pH changes, we found that recombinant APC trimer showed good absorbance and fluorescence stability at varying pH. The conformation of chromophores in recombinant APC trimer was relatively stable, between the range of pH 4-9, which was revealed by the stable absorption and fluorescence spectra properties of recombinant APC trimer. The trimeric structure of recombinant APC was maintained while local variations of protein peptides were also shown in response to the environmental disturbance. Beyond this pH range, secondary structure as well as overall conformation of recombinant APC dramatically changed, and the energy absorption and transfer ability were also disrupted. This work demonstrated that correct function in energy transfer was retained in the recombinant allophycocyanin trimer compared with nature allophycocyanin trimer.
中文摘要以基因重组别藻蓝蛋白三聚体为材料,利用原子力显微镜观察到了单分子重组别藻蓝蛋白三聚体的纳米级分辨率的三维图像(短轴约为1.5 nm,长轴约为15 nm的盘状颗粒),与通过透射电子显微镜和X射线晶体衍射得到的天然APC三聚体的圆盘状结构的尺寸十分接近.通过测量荧光发射光谱、吸光光谱和圆二色光谱,对自组装重组别藻蓝蛋白三聚体的正确组装进行了证实.获得了较宽pH范围内的紫外光区(190~250 nm)和可见光区(450~750 nm)圆二色谱数据,通过紫外光区圆二色谱解析,确定了a螺旋是重组别藻蓝蛋白三聚体二级结构的主要组成部分.可见光区圆二色谱解析为别藻蓝蛋白三聚体中存在藻蓝胆素激子耦合对,以及耦合体内以激子分裂进行能量传递提供了证据.本文结果进一步证实,与天然别藻蓝蛋白三聚体相比较,基因重组别藻蓝蛋白三聚体有正确的能量传递功能.
文章类型Article
收录类别EI; CSCD
语种中文
关键词[WOS]Plant Sciences
研究领域[WOS]Plant Sciences; 植物科学
CSCD记录号CSCD:6003393
引用统计
文献类型期刊论文
条目标识符http://ir.yic.ac.cnhttp://ir.yic.ac.cn/handle/133337/24307
专题海岸带生物学与生物资源利用重点实验室_海岸带生物学与生物资源保护实验室
作者单位1.中国科学院烟台海岸带研究所;
2.中国科学院大学;
3.鲁东大学农学院
推荐引用方式
GB/T 7714李文军,蒲洋,牛壮,等. 重组别藻蓝蛋白三聚体结构与功能[J]. 科学通报,2017,62(16):1699-1713.
APA李文军,蒲洋,牛壮,薛春岭,&秦松.(2017).重组别藻蓝蛋白三聚体结构与功能.科学通报,62(16),1699-1713.
MLA李文军,et al."重组别藻蓝蛋白三聚体结构与功能".科学通报 62.16(2017):1699-1713.
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