Mechanosensitive channels of Escherichia coli: the MscL gene, protein, and activities.
Mechanosensitive channels of Escherichia coli: the MscL gene, protein, and activities.
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DOI：
10.1146/annurev.physiol.59.1.633
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发表时间：
1997
影响因子：
18.2
通讯作者：
S. Sukharev;P. Blount;B. Martinac;C. Kung
中科院分区：
文献类型：
--
作者：
S. Sukharev;P. Blount;B. Martinac;C. Kung
虽然机械感觉反应是普遍存在的和多样的，在大多数情况下，机械感觉的分子基础仍然是神秘的MscL，大肠杆菌及其细菌同系物的大电导的机械敏感通道是第一个，目前唯一的通道分子显示直接感测膜的机械拉伸。为了响应通过脂质双层传递的张力，MscL将其打开概率增加了几个数量级。在本综述中，我们描述了这个最简单的mechanosensory分子的识别，克隆，和第一套生物物理和结构数据。我们在巨噬细胞中发现了2.5ns的机械敏感电导。coli原生质球。利用色谱法富集靶点，膜片钳法测定脂质体重构组分中的通道活性，我们鉴定了MscL蛋白并克隆了mscL基因。MscL包含136个氨基酸残基（15kDa），具有两个高度疏水区域，并且驻留在细菌的内膜中。PhoA融合实验表明，蛋白质跨越膜两次，两端在细胞质中。光谱技术表明它是高度螺旋的。MscL串联体的表达和共价交联表明活性通道复合物是同源六聚体。我们已经确定了几个残基，当删除或取代，影响通道动力学或机械敏感性。虽然发现时是独特的，但在革兰氏阴性和革兰氏阳性细菌中都发现了高度保守的MscL同源物，这表明它们在细菌中无处不在的重要性。
Although mechanosensory responses are ubiquitous and diverse, the molecular bases of mechanosensation in most cases remain mysterious MscL, a mechanosensitive channel of large conductance of Escherichia coli and its bacterial homologues are the first and currently only channel molecules shown to directly sense mechanical stretch of the membrane. In response to the tension conveyed via the lipid bilayer, MscL increases its open probability by several orders of magnitude. In the present review we describe the identification, cloning, and first sets of biophysical and structural data on this simplest mechanosensory molecule. We discovered a 2.5-ns mechanosensitive conductance in giant E. coli spheroplasts. Using chromatographies to enrich the target and patch clamp to assay the channel activity in liposome-reconstituted fractions, we identified the MscL protein and cloned the mscL gene. MscL comprises 136 amino acid residues (15 kDa), with two highly hydrophobic regions, and resides in the inner membrane of the bacterium. PhoA-fusion experiments indicate that the protein spans the membrane twice with both termini in the cytoplasm. Spectroscopic techniques show that it is highly helical. Expression of MscL tandems and covalent cross-linking suggest that the active channel complex is a homo-hexamer. We have identified several residues, which when deleted or substituted, affect channel kinetics or mechanosensitivity. Although unique when discovered, highly conserved MscL homologues in both gram-negative and gram-positive bacteria have been found, suggesting their ubiquitous importance among bacteria.