Tenzer Stefan, Hain Tobias, Berger Hendrik, Schild Hansjörg
Institute for Immunology, University of Mainz, Mainz, Germany.
Methods Mol Biol. 2013;960:1-14. doi: 10.1007/978-1-62703-218-6_1.
Proteasomes are the main cytosolic proteases responsible for generating peptides for antigen processing and presentation in the MHC (major histocompatibility complex) class-I pathway. Purified 20S and 26S proteasomes have been widely used to study both specificity and efficiency of antigen processing. Here, we describe the purification of active human 20S and 26S proteasomes from human erythrocytes by DEAE-ion exchange chromatography, ammonium sulfate precipitation, glycerol density gradient centrifugation, and Superose-6 size exclusion chromatography and their characterization using fluorogenic substrates and specific inhibitors.
蛋白酶体是主要的胞质蛋白酶，负责生成用于在主要组织相容性复合体（MHC）I类途径中进行抗原加工和呈递的肽段。纯化的20S和26S蛋白酶体已被广泛用于研究抗原加工的特异性和效率。在此，我们描述了通过DEAE离子交换色谱、硫酸铵沉淀、甘油密度梯度离心和Superose-6尺寸排阻色谱从人红细胞中纯化活性人20S和26S蛋白酶体，并使用荧光底物和特异性抑制剂对其进行表征。
