Structural analysis of the asparagine-linked glycan units of the ZP2 and ZP3 glycoproteins from mouse zona pellucida.
Structural analysis of the asparagine-linked glycan units of the ZP2 and ZP3 glycoproteins from mouse zona pellucida.
复制标题
小鼠透明带 ZP2 和 ZP3 糖蛋白的天冬酰胺连接聚糖单元的结构分析。
DOI：
10.1006/abbi.2000.2038
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发表时间：
2000
影响因子：
3.9
通讯作者：
Tulsiani,DR
中科院分区：
文献类型：
--
作者：
Tulsiani,DR
卵透明带（ZP）是哺乳动物卵细胞质膜外的一种糖萼，由三到四种糖蛋白组成。在小鼠中，ZP由三种糖蛋白组成，即ZP 1（200 kDa）、ZP 2（120 kDa）和ZP 3（83 kDa）。在该种属中的广泛研究已鉴定出精子的初级（mZP 3）和次级（mZP 2）结合位点。两种糖基化组分是高度糖基化的，含有N-连接和O-连接聚糖单元。为了表征N-连接聚糖单元，对mZP 2和mZP 3进行纯化，并通过N-聚糖酶彻底消化释放N-连接碳水化合物链。释放的低聚糖（OS）通过用NaB 3 H4还原进行放射性标记，并通过Bio-Gel P-4柱上的凝胶过滤进行分离。OS分离成几个峰，表明存在多种N-连接聚糖。有趣的是，从mZP 2和mZP 3分离的放射性峰非常不同，这一结果表明聚糖的定性和定量差异。分别合并mZP 2和mZP 3中存在的[3 H]标记聚糖，并通过系列凝集素色谱法进行分级。本报告中包含的实验证据强烈表明，mZP 3（而非mZP 2）含有末端非还原性α-半乳糖残基的聚乳糖胺聚糖。从固定化凝集素柱洗脱的mZP 3聚糖在用内切/外切糖水解酶消化之前或之后通过凝集素和尺寸柱色谱法进一步表征。数据显示存在多种OS，包括聚-N-乙酰基乳糖胺基、双-、三-和四触角复合型和高甘露糖型聚糖。总而言之，这些结果为mZP糖缀合物上存在的聚糖链的复杂性提供了额外的证据。
Zona pellucida (ZP), the extracellular glycocalyx that surrounds the mammalian egg plasma membrane, is a relatively simple structure consisting of three to four glycoproteins. In the mouse, the ZP is composed of three glycoproteins, namely ZP1 (200 kDa), ZP2 (120 kDa), and ZP3 (83 kDa). Extensive studies in this species have resulted in the identification of primary (mZP3) and secondary (mZP2) binding sites for spermatozoa. The two zona components are highly glycosylated containing N-linked and O-linked glycan units. In an attempt to characterize N-linked glycan units, mZP2 and mZP3 were purified and the N-linked carbohydrate chains were released by exhaustive digestion with N-glycanase. The released oligosaccharides (OSs) were radiolabeled by reduction with NaB3H4and resolved by gel filtration on a column of Bio-Gel P-4. The OSs separated into several peaks indicating the presence of a variety of N-linked glycans. Interestingly, the radioactive peaks resolved from mZP2 and mZP3 were quite different, a result suggesting qualitative and quantitative differences in the glycans. The [3H]-labeled glycans present in mZP2 and mZP3 were pooled separately and fractionated by serial lectin chromatography. Experimental evidence included in this report strongly suggests that mZP3 (but not mZP2) contains polylactosaminyl glycan with terminal, nonreducing α-galactosyl residues. The mZP3 glycans eluted from the immobilized lectin columns were further characterized by lectin and sizing column chromatography before or after digestion with endo-/exo-glycohydrolases. Data revealed the presence of a variety of OSs, including poly-N-acetyllactosaminyl, bi-, tri-, and tetraantennary complex-type, and high-mannose-type glycans. Taken together, these results provide additional evidence on the complex nature of the glycan chains present on mZP glycoconjugates.