Understanding ERAP molecular mechanism of MHC I antigen processing

摘要 在过去的十年中，I类抗原呈递途径的表征进展教导了 我们认为，抗原加工、肽转运、肽修整、肽选择和肽装载是 主要组织相容性提供的最佳肽库发展的关键事件 在一些实施方案中，免疫抑制剂可将MHC I类复合物（MHC I类分子）与细胞毒性T淋巴细胞（CTL）结合。这些事件影响了 产生构象稳定的MHC I/肽复合物所需的肽。证据 前提是MCH I肽库的总体质量的变化可以对CTL产生深远的影响。 应答因此，研究MHC I成熟过程中的基本生物学事件是至关重要的， 不仅是因为抗原加工和呈递的基本原理的进步，而且也是为了更好地 理解这些成熟事件如何调节免疫反应。 我们已经研究了肽装载和选择机制的分子和结构基础， MHC I已经超过20年了。最近，与抗原相关的内质网氨肽酶 ERAP 1、ERAP 2和ERAP 1，2异源二聚体（简称ERAP）已成为关键蛋白 用于影响MHC I肽库的形成。迄今为止，我们对ERAP如何发挥作用的理解， 然而，肽编辑器是模糊的。类似地，ERAP 1和ERAP 2多态性如何发挥作用的问题， 在慢性自身免疫性疾病中的作用，如HLA-B*27相关的强直性脊柱炎（AS），以及如何 ERAP 1和ERAP 2与疾病相关的MHC I分子相互作用，仍然不清楚。在本申请中， 我们将使用一种综合的方法来阐明ERAPs的功能和分子生物学的作用。 ERAP和MHC I分子之间的串扰。使用由ERAPs、MHC I类组成的无细胞系统， 分子，以及合成的和天然的N-末端延伸的肽，以及与生物化学， 分子和晶体学技术，我们将表征ERAPs对游离和MHC的功能， I-结合肽（目标#1）;阐明ERAP 1、HLA-B*27和AS之间的功能联系（目标#2）;以及 确定有和没有ERAP 1的HLA-B*0801/前体复合物的X射线晶体结构（目标#3）。 总的来说，我们的研究从根本上意义重大，因为它将提供新的知识，以更好地了解 ERAPs如何发挥作用并影响MHC I肽库的形成， ERAP 1与MHC分子间的相互作用 I分子。我们研究的医学意义是巨大的。

ABSTRACT Advances in characterization of the class I antigen presentation pathway over the last decade have taught us that antigen processing, peptide transport, peptide trimming, peptide selection, and peptide loading are critical events for the development of optimal peptide repertoires presented by major histocompatibility complex (MHC) class I molecules to cytotoxic T lymphocytes (CTLs). These events influence the tight binding of peptides needed to generate conformationally stable MHC I/peptide complexes. Evidence has been provided that changes in the overall quality of the MCH I peptide repertoire can have profound effects on CTL responses. Therefore, studies that characterize basic biological events in MHC I maturation are essential not only for advances in fundamental principles of antigen processing and presentation, but also to better comprehend how these maturation events modulates immune responses. We have studied the molecular and structural basis of mechanisms of peptide loading and selection by MHC I for more than 20 years. Recently, the endoplasmic reticulum-aminopeptidases associated with antigen processing (ERAP1, ERAP2, and ERAP1,2 heterodimer; referred to as ERAP) have emerged as key proteins for influencing formation of the MHC I peptide repertoire. To date, our understanding of how ERAP functions as a peptide editor is however obscure. Similarly, the question of how ERAP1 and ERAP2 polymorphisms play a role in chronic autoimmune diseases such as HLA-B*27-associated ankylosing spondylitis (AS), and how ERAP1 and ERAP2 interact with disease-associated MHC I molecules, still remain unclear. In this application, we will use a comprehensive approach to shed some light on the function of ERAPs and on the molecular crosstalk between ERAPs and MHC I molecules. Using a cell-free system composed of ERAPs, MHC class I molecules, and synthetic and natural N-terminally extended peptides, and in combination with biochemical, molecular, and crystallographic techniques, we will characterize the function of ERAPs towards free and MHC I-bound peptides (Aim #1); elucidate the functional links between ERAP1, HLA-B*27, and AS (Aim #2); and determine the x-ray crystal structure of an HLA-B*0801/precursor complex, with and without ERAP1 (Aim #3). Overall, our study is fundamentally significant because it will provide new knowledge to better understand how ERAPs function and influence formation of the MHC I peptide repertoire, novel insights into molecular events underlying the pathogenesis of AS, and will reveal the molecular interaction between ERAP1 and MHC I molecules. The medical relevance of our studies is immense.