Glycopeptides and Other Non-Natural Variants: Probes of Carbohydrate Function

描述(由申请人提供)：凝集素在免疫系统中发挥关键作用。树突状细胞上的凝集素参与抗原识别和内化，因此是免疫的关键介质。一种这样的凝集素DC-SIGN，它介导了有益的抗原摄取加工过程，可以被病原体所选择，以抑制免疫反应并促进其传播。例如，DC-SIGN可以与HIV结合并促进其传播。它还被证明可以抑制对结核分枝杆菌的免疫反应。由于HIV和结核分枝杆菌是全球范围内对人类健康构成最大威胁的两种疾病，因此阐明DC-SIGN功能的分子机制至关重要。这项拟议研究的长期目标是利用化学生物学阐明DC-SIGN如何在信号传递、抗原识别和抗原内化中发挥作用。对这些过程的深入了解可以帮助设计出提高疫苗效力或预防病原体感染的试剂。该项目的具体目标如下：(1)确定DC-SIGN的非碳水化合物、小分子配体；(2)开发跟踪DC-SIGN介导的抗原内化和树突状细胞迁移的化学成像策略；(3)开发新的化学方法来探索DC-SIGN介导的细菌-树突状细胞相互作用。第一个具体目标是提供一组新的DC-SIGN抑制剂。这些也可以作为研究使用这种凝集素的后果的工具。到目前为止，DC-SIGN配体是基于碳水化合物的，碳水化合物结合很弱，可以与其他树突状细胞表面凝集素相互作用。我们预计非碳水化合物配体将以更高的特异性和亲和力与DC-SIGN结合。在目标2中，我们将使用目标1中确定的配体来探索抗原与DC-SIGN相互作用的后果。DC-SIGN与多种不同类型的抗原(蛋白质、病毒、细菌和真菌)相互作用；因此，了解抗原结构如何影响内化和运输至关重要。我们将利用化学合成来改变DC-SIGN的配基结构，并将这些模型抗原赋予荧光报告基团进行成像。目的3探讨DC-SIGN参与如何影响生物体的树突状细胞内化和加工。为了研究这一过程，我们将对细菌进行化学修饰，使其显示特定的DC-SIGN配体。AIM 3中提出的研究结果可以深入了解DC-SIGN的作用，并为探索特定受体在宿主-病原体相互作用中的作用提供新的策略。关联性。碳水化合物结合蛋白DC-SIGN存在于树突状细胞表面，在免疫系统中发挥作用。除了在人类身上的正常功能外，DC-SIGN还可以被HIV等病毒用来促进它们的传播，也可以被一些细菌(如引起结核病的细菌)用来抑制免疫反应。由于DC-SIGN在促进导致全球最多死亡的两种疾病方面的作用，DC-SIGN如何工作以及如何控制它是重要的。本项目旨在鉴定DC-SIGN抑制剂，并利用与DC-SIGN结合的化合物来研究其功能。

DESCRIPTION (provided by applicant): Lectins play critical roles in the immune system. The lectins on dendritic cells participate in antigen recognition and internalization and thereby serve as critical mediators of immunity. One such lectin, DC-SIGN, which mediates the beneficial process of antigen uptake for processing, can be co-opted by pathogens to suppress immune responses and promote their dissemination. For example, DC-SIGN can bind to HIV and facilitate its dissemination. It also has been shown to suppress immune responses to Mycobacterium tuberculosis. Because HIV and Mycobacterium tuberculosis are the two most significant threats to human health worldwide, it is critical to elucidate the molecular mechanisms underlying the function of DC-SIGN. The long-term goals of the proposed research are to use chemical biology to illuminate how DC-SIGN functions in signaling, antigen recognition, and antigen internalization. Insight into these processes can facilitate the design of agents to enhance the effectiveness of vaccines or prevent pathogen infection. The specific aims of the project follow: (1) to identify non-carbohydrate, small-molecule ligands for DC- SIGN, (2) to develop chemical imaging strategies to follow DC-SIGN-mediated antigen internalization and dendritic cell migration, and (3) to develop new chemical approaches to explore bacteria-dendritic cell interactions mediated by DC-SIGN. The first specific aim focuses on providing a novel set of inhibitors of DC-SIGN. These also can serve as tools for exploring the consequences of engaging this lectin. The DC-SIGN ligands identified to date are based on carbohydrates, which bind weakly and can interact with other dendritic cell-surface lectins. We envision that non-carbohydrate-based ligands will bind to DC-SIGN with higher specificity and affinity. In Aim 2, we shall use the ligands identified in Aim 1 to explore the consequences of antigen interaction with DC-SIGN. DC-SIGN interacts with a variety of different types of antigens (proteins, viruses, bacteria, and fungi); therefore, it is critical to understand how antigen structure influences internalization and trafficking. We shall use chemical synthesis to vary DC-SIGN ligand structure and to endow these model antigens with fluorogenic reporter groups for imaging. Aim 3 addresses how dendritic cell internalization and processing of organisms is influenced by DC-SIGN engagement. To investigate this process, we shall chemically modify bacteria such that they display specific DC-SIGN ligands. The results of the investigations proposed in Aim 3 can provide insight into the role of DC-SIGN and offer new strategies to explore the role of specific receptors in host-pathogen interactions. Relevance. The carbohydrate-binding protein DC-SIGN is found on the surface of dendritic cells, where it functions in the immune system. In addition to its normal function in humans, DC-SIGN can be used by viruses, like HIV, to facilitate their dissemination, and by some bacteria, like those that cause tuberculosis, to suppress immune responses. Because of its role in facilitating the two diseases that cause the most deaths worldwide, how DC- SIGN works and how to control it is important. This project is aimed at identifying DC-SIGN inhibitors and using compounds that bind to DC-SIGN to study its function.