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首页> 外文期刊>The FEBS journal >Mutagenesis at the l-o interface impairs the cleavage of the dystroglycan precursor
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Mutagenesis at the l-o interface impairs the cleavage of the dystroglycan precursor

机译：在l-o界面上的诱变会破坏dystroglycan前体的裂解。

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The interaction between l-dystroglycan (l-DG) and o-dystroglycan (o-DG), the two constituent subunits of the adhesion complex dystroglycan, is crucial in maintaining the integrity of the dystrophin-glycoprotein complex. The importance of the l-o interface can be seen in the skeletal muscle of humans affected by severe conditions, such as Duchenne muscular dystrophy, where the l-o interaction can be secondarily weakened or completely lost, causing sarcolemmal instability and muscular necrosis. The reciprocal binding epitopes of the two subunits reside within the C-terminus of l-DG and the ectodomain of o-DG. As no ultimate structural data are yet available on the l-o interface, site-directed mutagenesis was used to identify which specific amino acids are involved in the interaction. A previous alanine-scanning analysis of the recombinant o-DG ectodomain allowed the identification of two phenylalanines important for l-DG binding, namely F692 and F718. In this article, similar experiments performed on the l-DG C-terminal domain pinpointed two residues, G563 and P565, as possible binding counterparts of the two o-DG phenylalanines. In 293-Ebna cells, the introduction of alanine residues instead of F692, F718, G563 and P565 prevented the cleavage of the DG precursor that liberates l- and o-DG, generating a pre-DG of about 160 kDa. This uncleaved pre-DG tetramutant is properly targeted at the cell membrane, is partially glycosylated and still binds laminin in pull-down assays. These data reinforce the notion that DG processing and its membrane targeting are two independent processes, and shed new light on the molecular mechanism that drives the maturation of the DG precursor. Structured digital abstract MINT-7214494: alpha DG (uniprotkb: Q62165) binds ( MI:0407) to beta DG (uniprotkb: Q62165) by solid phase assay ( MI:0892) MINT-7214516: laminin (uniprotkb: P19137) binds ( MI:0407) to beta DG (uniprotkb: Q62165) by pull down ( MI:0096)
机译：粘附复合物dystroglycan的两个组成亚基l-dystroglycan（l-DG）和o-dystroglycan（o-DG）之间的相互作用对于维持肌营养不良蛋白-糖蛋白复合物的完整性至关重要。 L-o界面的重要性可见于受严重状况影响的人的骨骼肌，例如Duchenne肌营养不良症，其中L-o相互作用可继而减弱或完全丧失，从而导致肌膜不稳定性和肌肉坏死。这两个亚基的相互结合表位位于1-DG的C末端和o-DG的胞外域内。由于在l-o界面上尚无最终的结构数据，因此使用定点诱变来鉴定相互作用中涉及哪些特定氨基酸。先前对重组o-DG胞外域的丙氨酸扫描分析允许鉴定出对I-DG结合很重要的两个苯基丙氨酸，即F692和F718。在本文中，对I-DG C末端结构域进行的类似实验指出了两个残基G563和P565，它们可能是两个o-DG苯丙氨酸的结合对应物。在293-Ebna细胞中，引入丙氨酸残基代替F692，F718，G563和P565阻止了释放l-和o-DG的DG前体的裂解，从而产生了约160 kDa的pre-DG。该未切割的pre-DG四突变体已正确靶向细胞膜，被部分糖基化，在下拉检测中仍与层粘连蛋白结合。这些数据强化了DG处理及其膜靶向是两个独立过程的观念，并为驱动DG前体成熟的分子机制提供了新的思路。结构化数字摘要MINT-7214494：αDG（uniprotkb：Q62165）通过固相测定（MI：0892）与βDG（uniprotkb：Q62165）结合（MI：0407）MINT-7214516：laminin（uniprotkb：P19137）结合（MI ：0407）下拉至Beta DG（uniprotkb：Q62165）（MI：0096）

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