Structural Characterization of a Heparan Sulfate Pentamer Interacting with LAR-Ig1-2.
Structural Characterization of a Heparan Sulfate Pentamer Interacting with LAR-Ig1-2.
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DOI：
10.1021/acs.biochem.8b00241
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发表时间：
2018-04-17
期刊：
影响因子：
2.9
通讯作者：
Prestegard JH
中科院分区：
文献类型：
--
作者：
Gao Q;Yang JY;Moremen KW;Flanagan JG;Prestegard JH
白细胞共同抗原相关蛋白（LAR）是IIa型受体蛋白酪氨酸磷酸酶（RPTPs）之一，在轴突生长和再生等生物过程中起着重要的信号转导作用。糖胺聚糖链，包括硫酸肝素（HS）和硫酸软骨素（CS），作为配体调节LAR信号传导。本文采用基于溶液的核磁共振方法，研究了LAR与HS五糖（GlcNS6S-GlcA-GlcNS3,6S-IdoA2S-GlcNS6S-OME, fondaparinux）相互作用的前两个免疫球蛋白结构域（Ig1-2）的结构特征。在研究过程中，我们扩展了一种分配策略，该策略适用于在哺乳动物细胞培养中表达的稀疏标记蛋白质，补充了一种类型的同位素富集氨基酸（在这种情况下为15n -赖氨酸），包括顺磁扰动到核磁共振。利用剩余偶极耦合数据，在溶液中验证了先前晶体结构中发现的LAR-Ig1-2的折叠双畴结构，并在对接程序HADDOCK中结合化学位移摄动对LAR-Ig1-2与fondaparinux的滴定、饱和转移差（STD）光谱和转移核Overhauser效应（trNOEs）来生成LAR-fondaparinux配合物的模型。这些模型通过后处理能量分析进一步分析，以确定关键的结合相互作用。除了深入了解IIa型RPTPs的配体相互作用机制以及CS和HS配体相反作用的起源外，这些结果可能有助于未来设计用于神经系统修复的治疗性化合物。
Leukocyte common antigen-related (LAR) protein is one of the type IIa receptor protein tyrosine phosphatases (RPTPs), which are important for signal transduction in biological processes including axon growth and regeneration. Glycosaminoglycan chains, including heparan sulfate (HS) and chondroitin sulfate (CS), act as ligands that regulate LAR signaling. Here, we report the structural characterization of the first two immunoglobulin domains (Ig1-2) of LAR interacting with an HS pentasaccharide (GlcNS6S-GlcA-GlcNS3,6S-IdoA2S-GlcNS6S-OME, fondaparinux) using multiple solution-based NMR methods. In the course of the study, we extended an assignment strategy useful for sparsely labeled proteins expressed in mammalian cell culture supplemented with a single type of isotopically enriched amino acid (15N-lysine in this case) by including paramagnetic perturbations to NMR resonances. The folded two domain structure for LAR-Ig1-2 seen in previous crystal structures has been validated in solution using residual dipolar coupling data, and a combination of chemical shift perturbation on titration of LAR-Ig1-2 with fondaparinux, saturation transfer difference (STD) spectra and transferred nuclear Overhauser effect (trNOEs) have been employed in the docking program, HADDOCK, to generate models for the LAR-fondaparinux complex. These models are further analyzed by post-processing energetic analysis to identify key binding interactions. In addition to providing insight into the ligand interaction mechanisms of type IIa RPTPs and the origin of opposing effects of CS and HS ligands, these results may assist in future design of therapeutic compounds for nervous system repair.
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影响因子：
15
作者：
Barthelmes, Katja;Reynolds, Anne M.;Peisach, Ezra;Jonker, Hendrik R. A.;DeNunzio, Nicholas J.;Allen, Karen N.;Imperiali, Barbara;Schwalbe, Harald
通讯作者：
Schwalbe, Harald
影响因子：
4.6
作者：
Hsieh PH;Thieker DF;Guerrini M;Woods RJ;Liu J
通讯作者：
Liu J
影响因子：
7.3
作者：
Coles CH;Jones EY;Aricescu AR
通讯作者：
Aricescu AR
影响因子：
4
作者：
Gao, Qi;Chen, Cheng-Yu;Prestegard, James H.
通讯作者：
Prestegard, James H.
DOI：
10.1007/978-1-4939-2343-4_26
发表时间：
2015-01-01
期刊：
Methods in molecular biology (Clifton, N.J.)
影响因子：
--
作者：
Hadden, Jodi A;Tessier, Matthew B;Woods, Robert J
通讯作者：
Woods, Robert J