Department of Biochemistry and Molecular Biology, Charbonneau Cancer Institute, Cumming School of Medicine, University of Calgary, Calgary, AB, Canada.
Methods Mol Biol. 2022;2444:227-241. doi: 10.1007/978-1-0716-2063-2_14.
With a predicted molecular mass of 469 kDa, expression of recombinant DNA-dependent protein kinase catalytic subunit (DNA-PKcs) is challenging. However, DNA-PKcs is relatively abundant in human cells, making it possible to purify the endogenous protein. Here we describe a method to purify DNA-PKcs and its binding partner Ku70/80 from HeLa cells and describe conditions for transfer of DNA-PKcs and other large polypeptides for immunoblotting.
预计重组 DNA 依赖性蛋白激酶催化亚基（DNA-PKcs）的分子质量为 469 kDa，其表达具有挑战性。然而，DNA-PKcs 在人细胞中相对丰富，因此有可能纯化内源性蛋白。本文描述了一种从 HeLa 细胞中纯化 DNA-PKcs 及其结合伴侣 Ku70/80 的方法，并描述了用于 DNA-PKcs 和其他大多肽进行免疫印迹转移的条件。
