Preparation and analysis of seven major, topographically defined fragments of band 3, the predominant transmembrane polypeptide of human erythrocyte membranes.
Preparation and analysis of seven major, topographically defined fragments of band 3, the predominant transmembrane polypeptide of human erythrocyte membranes.
复制标题
条带 3（人红细胞膜的主要跨膜多肽）的七个主要的、拓扑确定的片段的制备和分析。
DOI：
10.1021/bi00600a013
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发表时间：
1978
期刊：
影响因子：
2.9
通讯作者：
H. Köhler
中科院分区：
文献类型：
--
作者：
T. Steck;J. J. Koziarz;M. K. Singh;G. Reddy;H. Köhler
带3是一种分子量约为90000道尔顿的跨膜多肽，被认为能促进人红细胞膜阴离子的转运。以前的研究表明，这种蛋白质的消化，而仍然膜结合产生大的，地形定义，重叠的片段，占其全部或几乎全部的质量。我们现在已经纯化了这些片段中的七个，利用选择性膜溶解，凝胶过滤，和十二烷基硫酸钠中的制备凝胶电泳。氨基酸分析表明，来自带3的外表面和跨膜区域的片段明显疏水，而细胞质表面片段相对极性;这些组成数据平行的水溶液中的溶解度和膜协会的各种片段的模式。用糜蛋白酶消化完整细胞产生38000-道尔顿的外表面和55000-道尔顿的跨膜片段。它们的表观分子量和它们的组合物的总和approximated带3多肽，表明在细胞外面的单一位点的裂解。在iBand 3处的胰凝乳蛋白酶消化，一种~ 90 000-道尔顿糖蛋白，是人红细胞膜的主要多肽，占蛋白质质量的约25%（参见Steck，1974）。谱带3被认为参与了阴离子的易化扩散
Band 3 is the~ 90 000-daltonmembrane-span-ning polypeptide believed to facilitate anion transport in the human erythrocyte membrane. Previous studies have shown that digestion of this protein while still membrane-bound generates large, topographically defined, overlapping frag-ments which account for all or nearly all of its mass. We have now purified seven of these fragments, utilizing selective membrane solubilization, gel filtration, and preparative gel electrophoresis in sodium dodecyl sulfate. Amino acid analysis revealed that fragments derived from theouter surface and membrane-spanning regions of band 3 were distinctly hydro-phobic, while cytoplasmic surface segments were relatively polar; these compositional data parallel the solubility in aqueous solutions and the mode of membrane association of the various fragments. Digestion of intact cells with chymotrypsin generated a 38 000-dalton outer-surface and a 55 000-dalton transmembrane fragment. The sums of their apparent molecular weights and of their compositions ap-proximated the band 3 polypeptide, suggesting a single site of cleavage at the extracellular face. Chymotryptic digestion at iBand 3, an~ 90 000-dalton glycoprotein, is the predominant polypeptide of the human erythrocyte membrane, comprising approximately 25% of the protein mass (cf. Steck, 1974). Band 3 is believed to be involved in the facilitated diffusion of anions