Towards understanding the glycan code: next generation structural glycobiology for accurate description of protein-glycan complexes

理解聚糖代码：准确描述蛋白质-聚糖复合物的下一代结构糖生物学

• 批准号: BB/P010660/1
• 负责人: Jesus Angulo
• 金额: $ 49.22万
• 依托单位: University of East Anglia
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2017
• 资助国家: 英国
• 起止时间: 2017 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FP010660%2F1
• 关键词: Towards; understanding; glycan; code; next

The extraordinary properties of living systems emerge as a result of dynamic interactions between their biomolecular components (proteins, nucleic acid, carbohydrates or glycans, ...). Despite the complexity of a system of millions of molecular interactions, the whole network of interactions is highly regulated for healthy organisms, and alterations in the regulation processes underlie all diseases. In this network, different molecules interact with different strengths (affinities), where strong interacting partners produce stable biomolecular complexes and weak binding molecules produce transient assemblies. A well "orchestrated" system of protein-ligand interactions of very different affinities is indeed responsible for most of the regulation processes in living organisms. Strong complexes are formed when sustained biological signals are needed, whereas weak interactions are recalled when quick cellular responses are required after temporary stimuli (signal transduction, reversible cell-cell contacts, transient interactions in host/pathogen recognition, etc.). For a complete understanding of life processes, it is necessary to investigate both strong and weak protein-ligand interactions, which encourages the development of novel approaches to characterize the 3D molecular structures of weak protein complexes. Strong protein-ligand interactions have been extensively investigated and many biologically relevant 3D complexes have been determined, as their intrinsic stability makes them amenable to a number of analytical techniques. However, for weak protein interactions many conventional approaches fail or become unreliable. For example, X-ray crystallography, a very powerful structural technique, show limitations for weak interactions as: (i) obtaining crystals of the complexes including the ligand is difficult, and (ii) the typically poorly defined electronic density that describe the ligand in the binding pocket. NMR spectroscopy, one of the most powerful techniques to study intermolecular interactions, has demonstrated its extraordinary capability for the detection of weak protein-ligand interactions in solution, through the use of ligand-based experiments, like Saturation Transfer Difference (STD) NMR spectroscopy. However, the translation from these experiments to 3D structures is currently not straightforward. We have published some improvements in the set up of STD NMR experiments to determine protein-ligand affinities and study multiple binding modes of ligands in a protein binding pocket. The present proposal stems from recent results in our research group that allow us to propose that STD NMR spectroscopy can provide more structural information for weak interactions than the map of ligand contacts, or group epitope mapping (GEM). Here, we propose to obtain the orientation of the ligand in the binding pocket, a much needed piece of information. We plan to generate new experimental restraints to drive the structural calculations of the complexes, to get accurate 3D structures. Until now, these experimental restraints have remained unexplored. Among the most biologically relevant weak interactions, those of proteins with glycans are essential steps in many cell-cell communication processes, and key in the infectivity of microbial pathogens. In particular, influenza virus exploits this for initial cell recognition, attachment, and release of new virions. In a different strategy, HIV covers its surface with host glycans to evade the immunological response. Interestingly, new broad neutralizing antibodies (bNAb) are being discovered which are able to stop infection and are elicited against those carbohydrates in that glycan shield. In the context of our collaborations with Prof. Rob Field (Norwich) and Dr. Katie Doores (London), we will apply the novel STD NMR approaches to investigate the molecular recognition processes in human and avian influenza virus, and in immunologically active bNAbs against HIV.

生命系统的非凡特性是其生物分子成分（蛋白质、核酸、碳水化合物或聚糖等）之间动态相互作用的结果。尽管数以百万计的分子相互作用的系统是复杂的，但整个相互作用的网络对健康的生物体是高度调节的，调节过程的改变是所有疾病的基础。在这个网络中，不同的分子以不同的强度（亲和）相互作用，强相互作用的伙伴产生稳定的生物分子复合物，弱结合的分子产生瞬时组装。一个由不同亲和度的蛋白质-配体相互作用组成的“精心安排”的系统，确实对生物体的大部分调节过程负有责任。强复合物在需要持续的生物信号时形成，而弱相互作用则在临时刺激（信号转导、可逆的细胞-细胞接触、宿主/病原体识别中的短暂相互作用等）后需要快速的细胞反应时被唤醒。为了全面了解生命过程，有必要研究强和弱蛋白质-配体相互作用，这鼓励开发新的方法来表征弱蛋白质复合物的3D分子结构。强蛋白质-配体相互作用已被广泛研究，许多生物学相关的3D复合物已被确定，因为它们的内在稳定性使它们适用于许多分析技术。然而，对于弱蛋白质相互作用，许多传统方法失败或变得不可靠。例如，x射线晶体学，一种非常强大的结构技术，显示出弱相互作用的局限性：(i)获得包括配体在内的配合物的晶体是困难的，（ii）描述结合口袋中配体的典型的不明确的电子密度。核磁共振波谱是研究分子间相互作用最强大的技术之一，通过使用基于配体的实验，如饱和转移差（STD）核磁共振波谱，已经证明了它在检测溶液中弱蛋白质-配体相互作用方面的非凡能力。然而，从这些实验到3D结构的转换目前并不直截了当。我们发表了一些改进的STD NMR实验的设置，以确定蛋白质与配体的亲和力和研究配体在蛋白质结合口袋中的多种结合模式。目前的建议源于我们研究小组最近的结果，这些结果使我们能够提出STD核磁共振光谱可以提供比配体接触图或基团表位作图（GEM）更多的弱相互作用结构信息。在这里，我们建议获得配体在结合袋中的方向，这是一个非常需要的信息。我们计划产生新的实验约束来驱动复合物的结构计算，以获得精确的3D结构。到目前为止，这些实验限制仍未被探索。在最具生物学相关性的弱相互作用中，蛋白质与聚糖的相互作用是许多细胞-细胞通信过程的重要步骤，也是微生物病原体感染性的关键。特别是，流感病毒利用这一点进行初始细胞识别、附着和释放新的病毒粒子。在一种不同的策略中，HIV用宿主聚糖覆盖其表面以逃避免疫反应。有趣的是，新的广泛中和抗体（bNAb）正在被发现，它能够阻止感染，并被诱导对抗糖聚糖屏蔽中的碳水化合物。在我们与Rob Field教授（Norwich）和Katie Doores博士（London）合作的背景下，我们将应用新的STD NMR方法来研究人类和禽流感病毒的分子识别过程，以及抗HIV的免疫活性bNAbs。

项目成果

Molecular Recognition of Natural and Non-Natural Substrates by Cellodextrin Phosphorylase from Ruminiclostridium Thermocellum Investigated by NMR Spectroscopy.

• DOI: 10.1002/chem.202102039
• 发表时间: 2021-11-11
• 期刊: CHEMISTRY-A EUROPEAN JOURNAL
• 影响因子: 4.3
• 作者: Gabrielli, Valeria;Munoz-Garcia, Juan C.;Pergolizzi, Giulia;De Andrade, Peterson;Khimyak, Yaroslav Z.;Field, Robert A.;Angulo, Jesus
• 通讯作者: Angulo, Jesus

Uncovering a novel molecular mechanism for scavenging sialic acids in bacteria.

• DOI: 10.1074/jbc.ra120.014454
• 发表时间: 2020-10-02
• 期刊: The Journal of biological chemistry
• 作者: Bell A;Severi E;Lee M;Monaco S;Latousakis D;Angulo J;Thomas GH;Naismith JH;Juge N
• 通讯作者: Juge N

Spin Diffusion Transfer Difference (SDTD) NMR: An Advanced Method for the Characterisation of Water Structuration Within Particle Networks

自旋扩散传递差 (SDTD) NMR：一种表征粒子网络内水结构的先进方法

• DOI: 10.26434/chemrxiv.12770813.v1
• 发表时间: 2020
• 作者: Gabrielli V

NleB/SseK-catalyzed arginine-glycosylation and enteropathogen virulence are finely tuned by a single variable position contiguous to the catalytic machinery.

• DOI: 10.1039/d1sc04065k
• 发表时间: 2021-09-22
• 期刊: Chemical science
• 影响因子: 8.4
• 作者: García-García A;Hicks T;El Qaidi S;Zhu C;Hardwidge PR;Angulo J;Hurtado-Guerrero R
• 通讯作者: Hurtado-Guerrero R

Exploring the sequence-function space of microbial fucosidases

探索微生物岩藻糖苷酶的序列功能空间

• DOI: 10.21203/rs.3.rs-3101218/v1
• 发表时间: 2023
• 作者: Gascueña A