Characterization of the interaction of a recombinant soluble neuroligin-1 with neurexin-1β
Characterization of the interaction of a recombinant soluble neuroligin-1 with neurexin-1β
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DOI：
10.1074/jbc.m306803200
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发表时间：
2003-12-12
影响因子：
4.8
通讯作者：
Taylor, P
中科院分区：
文献类型：
--
作者：
Comoletti, D;Flynn, R;Taylor, P
神经连接素是α/β-水解酶折叠家族的蛋白质，被发现是突触后跨膜蛋白，其胞外结构域与突触前配偶体（neurexin家族的蛋白质）相关。为了表征神经连接素与neurexin-beta相互作用的分子基础，我们表达了五种可溶性和可输出形式的重组DNA来源的神经连接素-1，通过在其羧基末端附近的跨膜跨度之前截短蛋白质。通过沉降平衡分析，功能性神经配素-1的胞外结构域以二聚体的形式结合。通过表面等离子体共振，我们建立了可溶性神经连接素-1结合neurexin-1 β，但同源的α/β-水解酶折叠蛋白，乙酰胆碱酯酶，未能与neurexins。神经连接素-1在神经连接素家族中具有独特的N-连接糖基化模式，并且糖基化及其加工修饰神经连接素活性。蛋白质的不完全加工和酶促去除神经连接素-1的寡糖链或末端唾液酸增强其活性，而neurexin-1 β的去糖基化并没有改变其缔合能力。特别是，303位的N-连接糖基化似乎是改变与neurexin-1 β结合的主要决定因素。我们在这里表明，糖基化处理的神经连接素，除了mRNA剪接和基因选择，有助于特异性的neurexin-beta/neuroligin-1协会。
Neuroligins, proteins of the alpha/beta-hydrolase fold family, are found as postsynaptic transmembrane proteins whose extracellular domain associates with presynaptic partners, proteins of the neurexin family. To characterize the molecular basis of neuroligin interaction with neurexin-beta, we expressed five soluble and exportable forms of neuroligin-1 from recombinant DNA sources, by truncating the protein before the transmembrane span near its carboxyl terminus. The extracellular domain of functional neuroligin-1 associates as a dimer when analyzed by sedimentation equilibrium. By surface plasmon resonance, we established that soluble neuroligins-1 bind neurexin-1beta, but the homologous alpha/beta-hydrolase fold protein, acetylcholinesterase, failed to associate with the neurexins. Neuroligin-1 has a unique N-linked glycosylation pattern in the neuroligin family, and glycosylation and its processing modify neuroligin activity. Incomplete processing of the protein and enzymatic removal of the oligosaccharides chain or the terminal sialic acids from neuroligin-1 enhance its activity, whereas deglycosylation of neurexin-1beta did not alter its association capacity. In particular, the N-linked glycosylation at position 303 appears to be a major determinant in modifying the association with neurexin-1beta. We show here that glycosylation processing of neuroligin, in addition to mRNA splicing and gene selection, contributes to the specificity of the neurexin-beta/neuroligin-1 association.