Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY, USA.
Department of Microbiology, Immunology & Infection Diseases, University of Calgary, Calgary, AB, Canada.
Methods Mol Biol. 2025;2854:143-151. doi: 10.1007/978-1-0716-4108-8_15.
Protein lysine acetylation involved in the antiviral innate immunity contributes to the regulation of antiviral inflammation responses, including type 1 interferon production and interferon-stimulated gene expression. Thus, investigation of acetylated antiviral proteins is vital for the complete understanding of inflammatory responses to viral infections. Immunoprecipitation (IP) assay with anti-targeted-protein antibody or with acetyl-lysine affinity beads followed by immunoblot provides a classical way to determine the potential modified protein in the antiviral innate pathways, whereas mass spectrometry can be utilized to identify the accurate acetylation lysine residues or explore the acetyl-proteomics. We demonstrate here comprehensive methods of protein lysine acetylation determination in virus-infected macrophages and embryonic fibroblast cells or proteins-overexpressed HEK 293 T cells in the context of antiviral innate immunity.
蛋白赖氨酸乙酰化参与抗病毒先天免疫，有助于调节抗病毒炎症反应，包括 1 型干扰素的产生和干扰素刺激基因的表达。因此，研究乙酰化抗病毒蛋白对于全面了解病毒感染引起的炎症反应至关重要。用针对靶蛋白的抗体或乙酰赖氨酸亲和珠进行免疫沉淀（IP）检测，然后进行免疫印迹，这是一种经典的方法，可以确定抗病毒先天途径中潜在的修饰蛋白，而质谱可以用于鉴定准确的乙酰化赖氨酸残基或探索乙酰化蛋白质组学。在这里，我们展示了在抗病毒先天免疫背景下，检测病毒感染的巨噬细胞和胚胎成纤维细胞或过表达蛋白的 HEK 293T 细胞中蛋白赖氨酸乙酰化的综合方法。
