Physiological Chemistry Department, Genentech, South San Francisco, CA 94080, USA.
Biochemistry Department, West Virginia University, Morgantown, WV 26506, USA.
Cell. 2021 Aug 19;184(17):4480-4494.e15. doi: 10.1016/j.cell.2021.07.004. Epub 2021 Jul 27.
In neutrophils, nicotinamide adenine dinucleotide phosphate (NADPH) generated via the pentose phosphate pathway fuels NADPH oxidase NOX2 to produce reactive oxygen species for killing invading pathogens. However, excessive NOX2 activity can exacerbate inflammation, as in acute respiratory distress syndrome (ARDS). Here, we use two unbiased chemical proteomic strategies to show that small-molecule LDC7559, or a more potent designed analog NA-11, inhibits the NOX2-dependent oxidative burst in neutrophils by activating the glycolytic enzyme phosphofructokinase-1 liver type (PFKL) and dampening flux through the pentose phosphate pathway. Accordingly, neutrophils treated with NA-11 had reduced NOX2-dependent outputs, including neutrophil cell death (NETosis) and tissue damage. A high-resolution structure of PFKL confirmed binding of NA-11 to the AMP/ADP allosteric activation site and explained why NA-11 failed to agonize phosphofructokinase-1 platelet type (PFKP) or muscle type (PFKM). Thus, NA-11 represents a tool for selective activation of PFKL, the main phosphofructokinase-1 isoform expressed in immune cells.
在中性粒细胞中，通过磷酸戊糖途径产生的烟酰胺腺嘌呤二核苷酸磷酸（NADPH）为 NADPH 氧化酶 NOX2 提供燃料，以产生活性氧物质来杀死入侵的病原体。然而，NOX2 活性的过度增加会加剧炎症，如急性呼吸窘迫综合征（ARDS）。在这里，我们使用两种无偏化学蛋白质组学策略表明，小分子 LDC7559 或更有效的设计类似物 NA-11 通过激活糖酵解酶磷酸果糖激酶-1 肝型（PFKL）并抑制磷酸戊糖途径的通量来抑制中性粒细胞中依赖 NOX2 的氧化爆发。因此，用 NA-11 处理的中性粒细胞中依赖 NOX2 的产物减少，包括中性粒细胞细胞死亡（NETosis）和组织损伤。PFKL 的高分辨率结构证实了 NA-11 与 AMP/ADP 变构激活位点的结合，并解释了为什么 NA-11 不能激动磷酸果糖激酶-1 血小板型（PFKP）或肌肉型（PFKM）。因此，NA-11 代表了一种选择性激活主要在免疫细胞中表达的 PFKL 的工具。
