Kinetic and Structural Investigation of the Mechanism of Action of HLA-DO

描述（由申请人提供）：HLA-DO抗原呈递作用机制的动力学和结构研究对免疫系统功能和机体防御疾病的能力至关重要。为了触发免疫反应，蛋白质抗原被转化为短肽，并装载到主要组织相容性复合体（MHC） I类或II类分子上，在抗原呈递细胞表面呈递。这些肽- mhc复合物可以被T淋巴细胞上的抗原特异性受体识别，从而引发免疫反应。这个复杂的过程受到多种因素的调节，以识别病原体，同时在对抗疾病时防止对健康细胞的损害。这些多肽装载到MHC II类分子上是由MHC II类样蛋白人白细胞抗原(HLA)-DM催化的。HLA-DM对于产生稳定的肽- mhc复合物和改变细胞表面呈现的肽库至关重要。HLA-DO也是MHC类ll样蛋白，在某些细胞类型（如B淋巴细胞）中与HLA-DM相关。HLA-DO抑制HLA-DM功能的机制尚不清楚。通过调节HLA-DM， HLA-DO调节呈现的肽库。然而，对这些分子之间的相互作用知之甚少，并且由于来自天然来源的HLA-DO的有限可用性阻碍了研究。这一提议旨在确定HLA-DO作用于HLA-DM催化肽负载的机制。为此，将纯化可溶性功能活性HLA-DO蛋白，并采用生物化学和结构技术相结合的方法。利用荧光共振能量转移法研究HLA-DO对HLA-DM催化肽交换的步长/秒的影响。HLA-DO和HLA-DO/HLA-DM复合物的x射线晶体学将用于阐明抑制的结构基础。我们将研究HLA-DO抑制的pH依赖性和肽依赖性。研究HLA-DO的作用模式有助于了解疾病的发病机制，如自身免疫性疾病或癌症等疾病如何逃避免疫系统，以及开发特异性靶向疾病的免疫疗法。抗原呈递是机体免疫反应抵御病原体的重要步骤。本文研究了这一过程的分子机制。这种对身体如何检测病原体的理解对于对抗感染非常重要，也可以用于设计针对疾病的治疗方法。

DESCRIPTION (provided by applicant): Kinetic and Structural Investigation of the Mechanism of Action of HLA-DO Antigen presentation is critical to immune system function and to the body's ability to defend against disease. To trigger an immune response, protein antigens are converted to short peptides and loaded onto major histocompatibility complex (MHC) class I or class II molecules for presentation at the surface of antigen-presenting cells. These peptide-MHC complexes can be recognized by antigen-specific receptors on T lymphocytes to elicit an immune response. This complex process is regulated by a variety of factors to allow recognition of pathogens while also preventing damage to healthy cells when fighting off disease. The loading of these peptides on to MHC class II molecules is catalyzed by the MHC class ll-like protein human leukocyte antigen (HLA)-DM. HLA-DM is critical for the generation of stable peptide-MHC complexes and alters the repertoire of peptides presented on the cell surface. HLA-DO is also a MHC class ll-like protein which associates with HLA-DM in certain cell types, like B lymphocytes. HLA-DO has been shown to inhibit HLA-DM function by an unknown mechanism. Through regulation of HLA-DM, HLA-DO modulates the repertoire of presented peptides. However, little is known about the interaction between these molecules and studies have been hindered by the limited availability of HLA-DO from native sources. This proposal is aimed at determining the mechanism of HLA-DO action on HLA-DM catalyzed peptide loading. To this end, soluble functionally active HLA-DO protein will be purified and a combination of biochemical and structural techniques will be employed. The step/s of HLA-DM catalyzed peptide exchange which are affected by HLA-DO will be studied using a fluorescence resonance energy transfer assay. X-ray crystallography of HLA-DO and the HLA-DO/HLA-DM complex will be used to elucidate the structural basis for inhibition. Both the pH dependence and the peptide dependence of HLA-DO inhibition will be investigated. Studying the HLA-DO mode of action is beneficial for understanding the pathogenesis of disease, such as autoimmune disease or how diseases like cancer evade the immune system, and the development of immunotheraputics to specifically target diseases. Antigen presentation is an important step in the body's immune response to protect against pathogens. This work studies the molecular mechanisms of this process. This understanding of how the body detects pathogens is important for fighting infection and can also be utilized in designing theraputics to target diseases.