Graduate Institute of Biotechnology, National Chung-Hsing University, Taichung 402, Taiwan.
Plant Physiol Biochem. 2012 Dec;61:80-7. doi: 10.1016/j.plaphy.2012.09.008. Epub 2012 Sep 26.
Accumulation of oil bodies was successfully induced in a microalga, Chlorella sp., cultured in a nitrogen-limited medium. The oil bodies were initially assembled as many small entities (mostly 0.1-1 μm), and lately found as a major irregular compartment (>3 μm) occupying more than half of the cell space. Approximately, two thirds of oil bodies isolated from Chlorella cells were broken and formed a transparent oil layer on top of the milky compact layer of the remaining stable oil bodies after being washed with 0.1% triton X-100. The stable oil bodies mainly comprised triacylglycerols as examined by thin layer chromatography analysis and confirmed by both Nile red and BODIPY stainings. Integrity of these stable oil bodies was maintained via electronegative repulsion and steric hindrance possibly provided by their surface proteins. Immunological cross-recognition revealed that a major protein of 29 kDa, tentatively identified as caleosin, was exclusively present in Chlorella oil bodies. Mass spectrometric analysis showed that the putative caleosin possessed a trypic fragment of 13 residues matching to that of a hypothetical caleosin in Picea sitchensis. With the aid of a degenerate primer designed according to the tryptic peptide, a complete cDNA fragment encoding this putative caleosin was obtained by PCR. Phylogenetic tree analysis supports that Chlorella caleosin is the most primitive caleosin found in oil bodies to date.
油体在一种微藻（小球藻）中成功地在氮限制的培养基中积累。油体最初是许多小实体（大多为 0.1-1μm）组装而成，后来发现是一个主要的不规则隔室（>3μm），占据了细胞空间的一半以上。大约三分之二从小球藻细胞中分离出的油体在被 0.1%曲通 X-100 洗涤后会破裂，并在剩余稳定油体的乳白色致密层上形成透明的油层。薄层色谱分析和尼罗红和 BODIPY 染色证实，稳定的油体主要由三酰基甘油组成。这些稳定的油体可能通过表面蛋白提供的电负性排斥和空间位阻来保持完整性。免疫交叉识别表明，一种 29kDa 的主要蛋白质，暂定鉴定为钙卫蛋白，仅存在于小球藻油体中。质谱分析显示，该假定的钙卫蛋白具有与 Picea sitchensis 中假定的钙卫蛋白匹配的 13 个残基的胰蛋白酶片段。根据该胰蛋白酶肽设计的简并引物的帮助下，通过 PCR 获得了编码该假定钙卫蛋白的完整 cDNA 片段。系统发育树分析支持小球藻钙卫蛋白是迄今为止在油体中发现的最原始的钙卫蛋白。
