Isotopic replacement of pigments and a lipid in chlorosomes from Chlorobium limicola: characterization of the resultant chlorosomes.
Isotopic replacement of pigments and a lipid in chlorosomes from Chlorobium limicola: characterization of the resultant chlorosomes.
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DOI：
10.1021/bi602586g
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发表时间：
2007-05
期刊：
影响因子：
2.9
通讯作者：
Yoshinori Kakitani;Kenkichi Harada;T. Mizoguchi;Y. Koyama
中科院分区：
文献类型：
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作者：
Yoshinori Kakitani;Kenkichi Harada;T. Mizoguchi;Y. Koyama
色素，包括细菌叶绿素（BChl）c，类胡萝卜素，和痕量的BChl a连同脂质，单半乳糖基甘油二酯（MGDG），用氯仿/甲醇（1：1 v/v）从水悬浮液（50 mM Tris-HCl，pH 8.0）的chlorosomes从Chlorobium limicola提取;其他脂质和蛋白质留在水层中通过漏斗分离。通过吹扫N2气体干燥氯仿层，溶解在甲醇中，并快速注入水层以重新组装叶绿体。该技术已被开发用于用13 C-BChl c替代一半的固有12 C-BChl c以鉴定分子间的13 C-BChl c。13 C磁偶极相关峰（其被认为通过将13 C-BChl c浓度降低到一半而将其强度降低到四分之一），并通过固态NMR光谱确定棒状元件中BChl c聚集体的结构。同位素取代的绿质体的特征在于（1）通过蔗糖密度梯度离心、zeta电位测量、电子显微镜和动态光散射测量来确定绿质体的形态，（2）通过13 C NMR光谱、电子吸收和圆二色谱以及低角X射线衍射来确定杆状元件中的颜料组装，和（3）通过亚皮秒时间分辨吸收光谱法测定颜料组装体中的激发态动力学。结果表明，重组后的绿色体具有（1）相似但更长的形态结构，（2）几乎相同的色素组装在杆元件，和（3）基本上相同的激发态动力学的色素组装。
Pigments including bacteriochlorophyll (BChl) c, carotenoids, and a trace of BChl a together with a lipid, monogalactosyl diglyceride (MGDG), were extracted with chloroform/methanol (1:1 v/v) from an aqueous suspension (50 mM Tris-HCl, pH 8.0) of chlorosomes from Chlorobium limicola; other lipids and proteins were left behind in the aqueous layer by funnel separation. The chloroform layer was dried by purging N2 gas, dissolved in methanol, and rapidly injected into the aqueous layer to reassemble chlorosomes. This technique has been developed to replace one-half of the inherent 12C-BChl c by 13C-BChl c to identify the intermolecular 13C...13C magnetic dipole correlation peaks (that are supposed to reduce their intensities to one-fourth by reducing the 13C-BChl c concentration into one-half) and to determine the structure of BChl c aggregates in the rod elements by means of solid-state NMR spectroscopy. The isotopically replaced chlorosomes were characterized (1) by sucrose density gradient centrifugation, zeta potential measurement, electron microscopy, and dynamic light scattering measurement to determine the morphology of chlorosomes, (2) by 13C NMR spectroscopy, electronic absorption and circular dichroism spectroscopies, and low-angle X-ray diffraction to determine the pigment assembly in the rod elements, and (3) by subpicosecond time-resolved absorption spectroscopy to determine the excited-state dynamics in the pigment assembly. The results characterized the reassembled chlorosomes to have (1) similar but longer morphological structures, (2) almost the same pigment assembly in the rod elements, and (3) basically the same excited-state dynamics in the pigment assembly.