HYDROXYL RADICAL FOOTPRINTING OF DECTIN CARBOHYDRATE BINDING SURFACE

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 Dectin-1是一种模式识别受体，通过与真菌碳水化合物的相互作用，在宿主中性粒细胞对真菌病原体的反应中发挥重要作用。然而，碳水化合物-蛋白质结合表面目前尚不清楚。为此，我们表达并分离了Dectin-1和高亲和力的葡聚糖磷酸。为了确定碳水化合物的结合表面，我们使用了通过过氧化氢闪光光解的羟基足迹来区分标记蛋白质在存在和不存在高亲和力葡聚糖磷酸的情况下。修饰后，我们将确定每个氧化部位的修饰程度，并使用表观氧化速率的差异来确定氧化部位溶剂可及性的差异。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Dectin-1 is a pattern recognition receptor that is known to have an important role in host neutrophil response to fungal pathogens via interactions with fungal carbohydrates. However, the carbohydrate-protein binding surface is currently unknown. To this end, dectin-1 has been expressed and isolated, along with a high-affinity glucan phosphate. In order to determine the carbohydrate binding surface, we are using hydroxyl radical footprinting by flash photolysis of peroxide to differentially label the protein in the presence and absence of a high-affinity glucan phosphate. After modification, we will determine the extent of modification at each oxidation site, and use differences in apparent rates of oxidation to determine differences in solvent accessibility at the oxidation site.