Atomic and Specificity Details of Mucin 1 O-Glycosylation Process by Multiple Polypeptide GalNAc-Transferase Isoforms Unveiled by NMR and Molecular Modeling.
Atomic and Specificity Details of Mucin 1 O-Glycosylation Process by Multiple Polypeptide GalNAc-Transferase Isoforms Unveiled by NMR and Molecular Modeling.
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通过NMR和分子建模揭示的多个多肽GalNAc-转移酶异构体的粘蛋白1 O-糖基化过程的原子和特异性细节。
DOI：
10.1021/jacsau.1c00529
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发表时间：
2022-03-28
期刊：
影响因子：
8
通讯作者：
Marcelo F
中科院分区：
文献类型：
--
作者：
Coelho H;Rivas ML;Grosso AS;Diniz A;Soares CO;Francisco RA;Dias JS;Compañon I;Sun L;Narimatsu Y;Vakhrushev SY;Clausen H;Cabrita EJ;Jiménez-Barbero J;Corzana F;Hurtado-Guerrero R;Marcelo F
多肽GalNAc-转移酶（GalNAc-Ts）的大家族精确地控制GalNAc 0-聚糖如何添加到粘蛋白的串联重复区中（例如，MUC1）。然而，粘蛋白中O-聚糖的明确定义和聚集模式背后的结构特征知之甚少。在此背景下，在本文中，我们公开了通过NMR光谱学由分子建模方案辅助的GalNAc-T2/T3/T4同种型的MUCl 0-糖基化的全过程。通过使用MUC 1，四个串联重复结构域作为底物，我们证实了不同的GalNAc-Ts亚型的糖基化偏好，并强调了在添加第一个GalNAc残基后，凝集素结构域在糖基化位点选择中的重要性。在糖基化底物中，具有多个受体位点，凝集素结构域有助于将受体位点定向到催化结构域。我们的实验表明，在这个过程中，相邻的串联重复序列是关键的受体位点的进一步糖基化的GalNAc-T2/T4在凝集素辅助的方式。我们的研究还表明，在掺入GalNAc残基的过程中，肽骨架中的局部构象变化，这可能解释GalNAc-T2/T3/T4对MUC 1底物的精细特异性。有趣的是，我们假设MUC 1中PDTRP序列的特异性盐桥和反向γ-转角构象是GalNAc-T4对该区域特异性背后的主要结构基序。此外，细胞内分析显示GalNAc-T4同种型是体内使免疫原性表位PDTRP的Thr糖基化的唯一同种型，这突出了GalNAc-T4在该表位的糖基化中的相关性。最后，本文建立的NMR方法可以扩展到其他糖基转移酶，如C1 GalT 1和ST 6 GalNAc-I，以确定对复杂粘蛋白受体底物的特异性。
The large family of polypeptide GalNAc-transferases (GalNAc-Ts) controls with precision how GalNAc O-glycans are added in the tandem repeat regions of mucins (e.g., MUC1). However, the structural features behind the creation of well-defined and clustered patterns of O-glycans in mucins are poorly understood. In this context, herein, we disclose the full process of MUC1 O-glycosylation by GalNAc-T2/T3/T4 isoforms by NMR spectroscopy assisted by molecular modeling protocols. By using MUC1, with four tandem repeat domains as a substrate, we confirmed the glycosylation preferences of different GalNAc-Ts isoforms and highlighted the importance of the lectin domain in the glycosylation site selection after the addition of the first GalNAc residue. In a glycosylated substrate, with yet multiple acceptor sites, the lectin domain contributes to orientate acceptor sites to the catalytic domain. Our experiments suggest that during this process, neighboring tandem repeats are critical for further glycosylation of acceptor sites by GalNAc-T2/T4 in a lectin-assisted manner. Our studies also show local conformational changes in the peptide backbone during incorporation of GalNAc residues, which might explain GalNAc-T2/T3/T4 fine specificities toward the MUC1 substrate. Interestingly, we postulate that a specific salt-bridge and the inverse γ-turn conformation of the PDTRP sequence in MUC1 are the main structural motifs behind the GalNAc-T4 specificity toward this region. In addition, in-cell analysis shows that the GalNAc-T4 isoform is the only isoform glycosylating the Thr of the immunogenic epitope PDTRP in vivo, which highlights the relevance of GalNAc-T4 in the glycosylation of this epitope. Finally, the NMR methodology established herein can be extended to other glycosyltransferases, such as C1GalT1 and ST6GalNAc-I, to determine the specificity toward complex mucin acceptor substrates.
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影响因子：
16.6
作者：
de Las Rivas M;Lira-Navarrete E;Daniel EJP;Compañón I;Coelho H;Diniz A;Jiménez-Barbero J;Peregrina JM;Clausen H;Corzana F;Marcelo F;Jiménez-Osés G;Gerken TA;Hurtado-Guerrero R
通讯作者：
Hurtado-Guerrero R
影响因子：
7.7
作者：
Bagdonaite, Ieva;Pallesen, Emil M. H.;Wandall, Hans H.
通讯作者：
Wandall, Hans H.
DOI：
10.1073/pnas.1902211116
发表时间：
2019-10-08
影响因子：
11.1
作者：
Fernandez, Amy J.;Daniel, Earnest James Paul;Samara, Nadine L.
通讯作者：
Samara, Nadine L.
影响因子：
14.8
作者：
de Las Rivas M;Paul Daniel EJ;Narimatsu Y;Compañón I;Kato K;Hermosilla P;Thureau A;Ceballos-Laita L;Coelho H;Bernadó P;Marcelo F;Hansen L;Maeda R;Lostao A;Corzana F;Clausen H;Gerken TA;Hurtado-Guerrero R
通讯作者：
Hurtado-Guerrero R
影响因子：
15
作者：
Coelho, Helena;Matsushita, Talcahiko;Marcelo, Filipa
通讯作者：
Marcelo, Filipa