Membrane glycoproteins from spermatozoa: partial characterization of an integral Mr = approximately 24,000 molecule from rat spermatozoa that is glycosylated during epididymal maturation.
Membrane glycoproteins from spermatozoa: partial characterization of an integral Mr = approximately 24,000 molecule from rat spermatozoa that is glycosylated during epididymal maturation.
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来自精子的膜糖蛋白：来自大鼠精子的完整 Mr = 大约 24,000 个分子的部分表征，该分子在附睾成熟过程中被糖基化。
DOI：
10.1095/biolreprod34.5.925
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发表时间：
1986
影响因子：
3.6
通讯作者：
Baker,JB
中科院分区：
文献类型：
--
作者：
Hamilton,DW;Wenstrom,JC;Baker,JB
用半乳糖氧化酶-NaB[~(3~)]_4和NaO_4-~(2+)-NAB[~(3~)]_4分别标记细胞表面半乳糖和唾液酸部分的大鼠附睾尾精子。经40 mM辛基-β-D-吡喃葡萄糖苷提取后，十二烷基硫酸钠-聚丙烯酰胺凝胶电泳显示，在11.2%、14%和16.8%的管状凝胶中，在Mr=∿24000处有一个单一的放射性峰迁移。相同的OBG提取物在5.6%和14%凝胶上的SDS-PAGE表明，该分子与其他报道的分子相同，分子量从32,000到37,000不等。用8M尿素和6M盐酸胍提取的标记分子量分别是OBG法的30%和50%。然而，当标记精子在其他条件下(在不同的温度和提取时间下，在pH 8，pH 3，0.1-3MNaC l[在pH 7.2]，5 mM乙二胺四乙酸，20-200 mM二硫苏糖醇或20-200 mMβ，在不同的温度和提取时间下)处理时，标记分子的萃取量不到OBG的1%。如Sephadex G-100层析和蔗糖密度梯度离心法所示，该分子在含水缓冲液中聚集。电荷转移电泳法分析表明，该分子含有一个暴露的疏水结构域(S)。温和的胰酶处理释放了所有标记的碳水化合物，大部分标记附着在MR=10,000的片段上。这些数据支持这一假设，即该分子是一个完整的膜糖蛋白，并表明它只部分埋藏在质膜的脂基质中。
Spermatozoa from the rat cauda epididymidis were treated with either the galactose oxidase-NaB[3H]4or the NaIO4-NaB [3H]4technique to label cell surface moieties of galactose and sialic acid, respectively. Following extraction with 40 mM octyl-β-D-glucopyranoside (OBG), electrophoresis in sodium dodecylsulfate-polyacrylamide gels (SDS-PAGE) revealed a single radioactive peak migrating at Mr= ∿ 24,000 in 11.2%, 14% and 16.8% tube gels. SDS-PAGE of the same OBG extract on 5.6% and 14% gels showed that this molecule was the same as that reported elsewhere, having a molecular weight varying from 32,000 to 37,000. The amount of labeled molecule extracted with 8 M urea or with 6 M guanidine-HCl was 30 and 50%, respectively, of that achieved with OBG. However, when labeled sperm were treated under other conditions (at pH 8, pH 3, 0.1–3 M NaCl [at pH 7.2], 5 mM ethylenediaminetetraacetic acid [EDTA], 20–200 mM dithiothreitol or 20–200 mM βmercaptoethanol, at varying temperatures and extraction times), the amount of labeled molecule extracted was less than 1% of that obtained with OBG. The molecule aggregates in aqueous buffers, as shown by chromatography using Sephadex G-100 and by centrifugation through a sucrose density gradient. Analysis by charge-shift electrophoresis suggested that the molecule contains an exposed hydrophobic domain(s). Mild trypsin treatment released all the labeled carbohydrate with the majority of the label attached to a Mr= 10,000 fragment. These data support the hypothesis that the molecule is an integral membrane glycoprotein, and suggest that it is only partially buried in the lipid matrix of the plasma membrane.