Structural and Mechanistic Insights into the Catalytic-Domain-Mediated Short-Range Glycosylation Preferences of GalNAc-T4.
Structural and Mechanistic Insights into the Catalytic-Domain-Mediated Short-Range Glycosylation Preferences of GalNAc-T4.
复制标题
DOI：
10.1021/acscentsci.8b00488
复制
发表时间：
2018-09-26
影响因子：
18.2
通讯作者：
Hurtado-Guerrero R
中科院分区：
文献类型：
--
作者：
de Las Rivas M;Paul Daniel EJ;Coelho H;Lira-Navarrete E;Raich L;Compañón I;Diniz A;Lagartera L;Jiménez-Barbero J;Clausen H;Rovira C;Marcelo F;Corzana F;Gerken TA;Hurtado-Guerrero R
粘蛋白型O-糖基化是由一系列多肽GalNAc-转移酶(GalNAc-ts)启动的，GalNAc-ts是一种包含高尔基体、腔催化和凝集素结构域的II型跨膜蛋白，它们通过一个灵活的连接子连接在一起。几种GalNAc-ts，包括GalNAc-T4，都表现出长程和短程糖基化前的特异性，分别由它们的凝集素和催化结构域控制。虽然凝集素结构域依赖的糖基化的机制是众所周知的，但糖肽的催化结构域依赖的糖基化的分子基础还不清楚。在这里，我们报道了GalNAc-T4与二糖肽GAT*GAGAGAGT*TPGPG(包含两个α-GalNAc糖基化的Thr(T*)、PxP基序和一个“裸露的”Thr受体位点)结合的晶体结构，描述了它的催化结构域糖肽GalNAc结合部位。对野生型和GalNAc结合位点突变酶的动力学研究表明，凝集素结构域的GalNAc结合活性高于催化结构域的GalNAc结合活性，这些活性可以独立消除。令人惊讶的是，从凝集素结构域伸出的柔性环被发现对于催化结构域的最佳活性是必不可少的。这项工作为GalNAc-T的短程糖基化偏好提供了第一个结构基础。首次表征和可视化了GalNAc-T的催化域糖肽GalNAc结合部位，揭示了GalNAc-T4‘S邻近O-糖基化活性的分子起源。
Mucin-type O-glycosylation is initiated by a family of polypeptide GalNAc-transferases (GalNAc-Ts) which are type-II transmembrane proteins that contain Golgi luminal catalytic and lectin domains that are connected by a flexible linker. Several GalNAc-Ts, including GalNAc-T4, show both long-range and short-range prior glycosylation specificity, governed by their lectin and catalytic domains, respectively. While the mechanism of the lectin-domain-dependent glycosylation is well-known, the molecular basis for the catalytic-domain-dependent glycosylation of glycopeptides is unclear. Herein, we report the crystal structure of GalNAc-T4 bound to the diglycopeptide GAT*GAGAGAGT*TPGPG (containing two α-GalNAc glycosylated Thr (T*), the PXP motif and a “naked” Thr acceptor site) that describes its catalytic domain glycopeptide GalNAc binding site. Kinetic studies of wild-type and GalNAc binding site mutant enzymes show the lectin domain GalNAc binding activity dominates over the catalytic domain GalNAc binding activity and that these activities can be independently eliminated. Surprisingly, a flexible loop protruding from the lectin domain was found essential for the optimal activity of the catalytic domain. This work provides the first structural basis for the short-range glycosylation preferences of a GalNAc-T. First characterization and visualization of the catalytic domain glycopeptide GalNAc binding site of a GalNAc-T revealing the molecular origins for GalNAc-T4’s neighboring O-glycosylation activity.
登录
查看更多内容
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：
16.6
作者：
Lira-Navarrete, Erandi;de las Rivas, Matilde;Companon, Ismael;Carmen Pallares, Maria;Kong, Yun;Iglesias-Fernandez, Javier;Bernardes, Goncalo J. L.;Peregrina, Jesus M.;Rovira, Carme;Bernado, Pau;Bruscolini, Pierpaolo;Clausen, Henrik;Lostao, Anabel;Corzana, Francisco;Hurtado-Guerrero, Ramon
通讯作者：
Hurtado-Guerrero, Ramon
影响因子：
2.9
作者：
Dam, Tarun K.;Gerken, Thomas A.;Brewer, C. Fred
通讯作者：
Brewer, C. Fred
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
DOI：
10.1073/pnas.1511175112
发表时间：
2015-11-24
影响因子：
11.1
作者：
Goth, Christoffer K.;Halim, Adnan;Schjoldager, Katrine T. -B. G.
通讯作者：
Schjoldager, Katrine T. -B. G.