The involvement of O-linked oligosaccharide chains of the sea urchin egg receptor for sperm in fertilization.
The involvement of O-linked oligosaccharide chains of the sea urchin egg receptor for sperm in fertilization.
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海胆卵子受体的 O-连接寡糖链参与受精。
DOI：
10.1093/glycob/5.1.11
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发表时间：
1995
期刊：
影响因子：
4.3
通讯作者：
Lennarz,WJ
中科院分区：
文献类型：
--
作者：
Dhume,ST;Lennarz,WJ
最近对海胆卵子精子受体的研究对其进行了测序，并证明它是一种 350 kDa 的糖蛋白。在当前的研究中，N-和O-连接的寡糖链从伴刀豆球蛋白A-琼脂糖上分级的蛋白质中裂解出来。通过阴离子交换色谱法进一步分离未与凝集素结合的推定的 O-连接寡糖链。使用竞争生物测定法测量这些寡糖链抑制受精的能力，发现 N 连接链缺乏抑制活性。相反，抑制活性局限于 O-连接链，带最高电荷的硫酸化链显示出最高的抑制活性。通过还原标记生物活性寡糖并分析其与精子的结合。结合测定的结果与受精生物测定相结合表明寡糖通过与顶体反应的精子结合来抑制受精。与完整受体和仅含有受体胞外结构域的重组糖蛋白不同，生物活性寡糖在受精生物测定中缺乏物种特异性。由于先前的工作表明重组蛋白特异性抑制受精物种并与顶体反应的精子结合，因此提出了精卵相互作用的两步模型。假设第一步是受体的硫酸化 O-连接寡糖链与精子之间的低亲和力离子相互作用，这种相互作用不具有物种特异性。随后精子与受体多肽链上的一个或多个结合位点发生高亲和力、物种特异性相互作用。
Recent investigations on the sea urchin egg receptor for sperm have led to its sequencing and the demonstration that it is a 350 kDa glycoprotein. In the current study, the N- and O-linked oligosaccharide chains were cleaved from the protein fractionated on concanavalin A-agarose. The putative O-linked oligosaccharide chains that did not bind to the lectin were further fractionated by anion-exchange chromatography. Using a competition bioassay that measured the ability of these oligosaccharide chains to inhibit fertilization, it was found that the N-linked chains were devoid of inhibitory activity. Rather, the inhibitory activity was localized to the O-linked chains, with the most highly charged, sulphated chains showing the highest inhibitory activity. The bioactive oligosaccharides were labelled by reduction and assayed for binding to sperm. The results of the binding assay, coupled with the fertilization bioassay, indicate that the oligosaccharides inhibit fertilization by binding to acrosome-reacted sperm. The bioactive oligosaccharide lacked species specificity in fertilization bioassays, unlike the intact receptor and a recombinant aglyco protein containing only the extracellular domain of the receptor. Since previous work showed that the recombinant protein inhibits fertilization species specifically and binds to acrosome-reacted sperm, a two-step model of sperm-egg interaction is proposed. The first step is postulated to be a low-affinity ionic interaction of the sulphated O-linked oligosaccharide chains of the receptor with sperm that is not species specific. This is followed by a high affinity, species-specific interaction of the sperm with one or more binding sits on the polypeptide chain of the receptor.