Relevance of serum mannosidase resistance to the specificity of oligomannose-targeting antibodies

项目摘要 修饰HIV包膜糖蛋白刺突（Env）的聚糖名义上被归类为“自身” 抗原，因为它们由哺乳动物细胞的糖基化机制赋予。然而，在这方面， 在gp 120上发现的低聚甘露糖型聚糖大部分聚集成一个小块，形成一个“非自身”的小块。 架构这种寡聚甘露糖贴片很容易被广泛中和的抗HIV抗体识别。 抗体（bnAbs）。鉴于bnAb对寡聚甘露糖斑块的效力和广度， 有兴趣开发免疫策略来诱导类似的bnAb。 该项目旨在填补理解寡聚甘露糖特异性bnAbs引发障碍的空白。 我们和其他人最近报道了在哺乳动物血清中发现的可溶性甘露糖苷酶， 寡甘露糖苷在糖缀合物上的应用，所述糖缀合物被设计为在 次免疫这种修剪可以解释为什么用寡甘露糖苷糖缀合物免疫 通常产生与寡聚甘露糖的亚结构结合良好但与寡聚甘露糖的亚结构结合良好的抗体。 对Env上呈现的全尺寸寡甘露糖的亲合力显著较低;后者代表结合 靶向寡聚甘露糖斑块的bnAb的方式。 在本申请中，我们建议确定血清甘露糖苷酶活性的相关程度 低聚甘露糖特异性抗体应答的质量。具体来说，我们打算建立一个 甘露糖苷酶抗性糖苷，因此代表对甘露糖苷酶的最大可能抗性 修剪，并确定随后的抗体反应的特异性相比，其甘露糖苷酶- 敏感的反面该项目将利用我们已发表的工作的细菌启发模仿 寡甘露糖，作为一种新糖缀合物，激发适度水平的HIV中和抗体。我们将 研究多克隆血清水平和单克隆水平的抗体应答;后者 如果有的话，应当揭示引发的抗体和现有bnAb之间的相似性。 总之，我们希望这个项目的结果能够揭示激发抗体的障碍， 以结合HIV Env上高度脆弱的寡聚甘露糖贴片。虽然很难引出 寡聚甘露糖特异性bnAb通常被认为源于B细胞耐受，我们的初步研究 提供血清甘露糖苷酶至少造成额外阻碍的证据。

PROJECT SUMMARY The glycans that decorate the HIV envelope glycoprotein spike (Env) are categorized nominally as ‘self’ antigens given that they are imparted by the mammalian cell’s glycosylation machinery. However, glycans of the oligomannose-type found on gp120 are mostly clustered into a patch, creating a ‘non-self’ architecture. This oligomannose patch is vulnerable to recognition by anti-HIV broadly neutralizing antibodies (bnAbs). Given the potency and breadth of bnAbs to the oligomannose patch, there is great interest in developing immunization strategies to elicit similar bnAbs. This project aims to fill a gap in understanding barriers to the elicitation of oligomannose-specific bnAbs. We and others have recently reported on a soluble mannosidase found in mammalian serum that trims oligomannosides on glycoconjugates designed to elicit oligomannose-specific antibodies upon immunization. Such trimming would explain why immunizing with oligomannosidic glycoconjugates typically yields antibodies that bind well to substructures of oligomannose but that bind with substantially lower avidity to full-sized oligomannose as presented on Env; the latter typifies the binding manner of bnAbs that target the oligomannose patch. In this application, we propose to determine the extent to which serum mannosidase activity is relevant to the quality of oligomannose-specific antibody responses. Specifically, we intend to construct a mannosidase-resistant glycoside, thus representing the greatest possible resistance to mannosidase trimming, and determine the specificity of the ensuing antibody response compared to its mannosidase- sensitive opposite. The project will take advantage of our published work of a bacterially inspired mimic of oligomannose that, as a neoglycoconjugate, elicits modest levels of HIV-neutralizing antibodies. We will investigate antibody responses at the polyclonal serum level as well as the monoclonal level; the latter should reveal similarities, if any, between the elicited antibodies and existing bnAbs. In sum, we expect the results of this project to shed light on barriers to evoking antibodies with capacity to bind the highly vulnerable oligomannose patch on HIV Env. Although the difficulty in eliciting oligomannose-specific bnAbs is often assumed to stem from B cell tolerance, our preliminary studies provide evidence for serum mannosidase posing at least an additional hindrance.