Biochemical studies of the excitable membrane of Paramecium tetraurelia. III. Proteins of cilia and ciliary membranes.
Biochemical studies of the excitable membrane of Paramecium tetraurelia. III. Proteins of cilia and ciliary membranes.
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DOI：
10.1083/jcb.84.3.717
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发表时间：
1980-03
期刊：
影响因子：
--
通讯作者：
Nelson DL
中科院分区：
文献类型：
--
作者：
Adoutte A;Ramanathan R;Lewis RM;Dute RR;Ling KY;Kung C;Nelson DL
作为野生型草履虫及其行为突变体膜激发的生化分析的第一步，我们确定了野生型细胞纤毛膜的蛋白质组成。改进了纤毛和纤毛膜泡的分离技术。在不使用洗涤剂的情况下，获得了高纯度和完整性的膜。用电子显微镜对各组分进行了表征，并用SDS-聚丙烯酰胺凝胶电泳法和等电聚焦对整个纤毛、轴丝和纤毛膜囊泡的蛋白质进行了一维和二维的分离。组分的蛋白质图谱和EM外观具有高度的重复性。在分离的纤毛中存在200多个多肽，其中大部分被回收到轴丝部分。毛囊有时作为纤毛制剂中的一种次要污染物，由一组非常不同的30多个多肽组成，摩尔重量在11,000-19,000之间。膜小泡含有多达70个摩尔质量为15,000-250,000的多肽。主要的囊泡种类是一种高分子量的蛋白质(“固定化抗原”)和一组摩尔质量接近或接近40,000的酸性蛋白质。这些和其他几种膜蛋白在轴丝部分特异性减少或完全缺失。微管蛋白是轴丝的主要物种，仅以微量形式存在于离体囊泡中；用本文所述方法制备的四膜虫纤毛膜也是如此。指数生长期细胞的囊泡中几乎不存在分子质量为31,000的蛋白，等电点为6.8，但在稳定期较早出现，与细胞的交配反应性有很好的相关性。这种详细的特性将为比较野生型和行为突变体的纤毛蛋白以及分析膜蛋白的形态和功能提供基础。这也将有助于未来对毛囊和交配反应的研究。
As a first step in the biochemical analysis of membrane excitation in wild-type Paramecium and its behavioral mutants we have defined the protein composition of the ciliary membrane of wild-type cells. The techniques for the isolation of cilia and ciliary membrane vesicles were refined. Membranes of high purity and integrity were obtained without the use of detergents. The fractions were characterized by electron microscopy, and the proteins of whole cilia, axonemes, and ciliary membrane vesicles were resolved by SDS polyacrylamide gel electrophoresis and isoelectric focusing in one and two dimensions. Protein patterns and EM appearance of the fractions were highly reproducible. Over 200 polypeptides were present in isolated cilia, most of which were recovered in the axonemal fraction. Trichocysts, which were sometimes present as a minor contaminant in ciliary preparations, were composed of a very distinct set of over 30 polypeptides of mol wt 11,000--19,000. Membrane vesicles contained up to 70 polypeptides of mol wt 15,000--250,000. The major vesicle species were a high molecular weight protein (the "immobilization antigen") and a group of acidic proteins with mol wt similar to or approximately 40,000. These and several other membrane proteins were specifically decreased or totally absent in the axoneme fraction. Tubulin, the major axonemal species, occurred only in trace amounts in isolated vesicles; the same was true for Tetrahymena ciliary membranes prepared by the methods described in this paper. A protein of mol wt 31,000, pI 6.8, was virtually absent in vesicles prepared from cells in exponential growth phase, but became prominent early in stationary phase in good correlation with cellular mating reactivity. This detailed characterization will provide the basis for comparison of the ciliary proteins of wild-type and behavioral mutants and for analysis of topography and function of membrane proteins. It will also be useful in future studies of trichocysts and mating reactions.
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DOI：
10.1083/jcb.78.2.451
发表时间：
1978-08
期刊：
The Journal of cell biology
影响因子：
--
作者：
Dute R;Kung C
通讯作者：
Kung C
DOI：
10.1083/jcb.69.1.126
发表时间：
1976-04
期刊：
The Journal of cell biology
影响因子：
--
作者：
Beisson J;Lefort-Tran M;Pouphile M;Rossignol M;Satir B
通讯作者：
Satir B
影响因子：
5.5
作者：
DUNLAP, K
通讯作者：
DUNLAP, K
DOI：
10.1016/0005-2736(79)90205-0
发表时间：
1979-01-01
期刊：
BIOCHIMICA ET BIOPHYSICA ACTA
影响因子：
--
作者：
ANDREWS, D;NELSON, DL
通讯作者：
NELSON, DL
影响因子：
7.8
作者：
HANSMA, HG
通讯作者：
HANSMA, HG