Morphology of isolated Gli349, a leg protein responsible for Mycoplasma mobile gliding via glass binding, revealed by rotary shadowing electron microscopy
Morphology of isolated Gli349, a leg protein responsible for Mycoplasma mobile gliding via glass binding, revealed by rotary shadowing electron microscopy
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DOI：
10.1128/jb.188.8.2821-2828.2006
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发表时间：
2006-04-01
影响因子：
3.2
通讯作者：
Miyata, M
中科院分区：
文献类型：
--
作者：
Adan-Kubo, J;Uenoyama, A;Miyata, M
几种支原体依靠一种未知的机制，在细胞极处的膜突起方向滑过固体表面。我们最近对移动支原体（Mycoplasma mobile）这一速度最快的物种的研究表明，一种349 kda的蛋白质Gli349位于被称为颈部的膜突起的底部，形成了从颈部伸出来的腿，并通过ATP水解的能量反复结合和释放固体表面来推动细胞。本文从支原体细胞中分离出Gli349蛋白，并对其结构进行了分析。凝胶过滤分析表明，分离得到的Gli349蛋白为单体。旋转阴影电子显微镜显示，分子结构类似于音乐中八分音符的符号。它包含一个椭圆形的脚长14纳米的轴。从这只脚延伸出三根杆，依次为43、20和20纳米。连接第一杆和第二杆的铰链是灵活的，而下一个铰链在其角度上有明显的偏好，接近90度。分子图像显示，一个单克隆抗体可以结合到距离N端总肽长度的三分之一的位置，并结合到距离足端三分之二的位置，表明足端对应于c端区域。将氨基酸序列分配到分子图像中，并讨论了分子在滑动机制中的拓扑结构。
Several species of mycoplasmas rely on an unknown mechanism to glide across solid surfaces in the direction of a membrane protrusion at the cell pole. Our recent studies on the fastest species, Mycoplasma mobile, suggested that a 349-kDa protein, Gli349, localized at the base of the membrane protrusion called the neck, forms legs that stick out from the neck and propel the cell by repeatedly binding to and releasing from a solid surface, based on the energy of ATP hydrolysis. Here, the Gli349 protein was isolated from mycoplasma cells and its structure was analyzed. Gel filtration analysis showed that the isolated Gli349 protein is monomeric. Rotary shadowing electron microscopy revealed that the molecular structure resembles the symbol for an eighth note in music. It contains an oval foot 14 nm long in axis. From this foot extend three rods in tandem of 43, 20, and 20 nm, in that order. The hinge connecting the first and second rods is flexible, while the next hinge has a distinct preference in its angle, near 90 degrees. Molecular images revealed that a monoclonal antibody that can bind to the position at one-third of the total peptide length from the N terminus bound to a position two-thirds from the foot end, suggesting that the foot corresponds to the C-terminal region. The amino acid sequence was assigned to the molecular image, and the topology of the molecule in the gliding machinery is discussed.