Purification of DNA-Dependent Protein Kinase Catalytic Subunit (DNA-PKcs) from HeLa Cells

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 a...

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Bibliographic Details
Published inMethods in molecular biology (Clifton, N.J.) Vol. 2444; p. 227
Main Authors Lee, Linda, Yu, Yaping, Lees-Miller, Susan P
Format Journal Article
LanguageEnglish
Published United States 2022
Subjects
Catalytic Domain
DNA - chemistry
DNA-Activated Protein Kinase - chemistry
DNA-Activated Protein Kinase - genetics
DNA-Activated Protein Kinase - metabolism
DNA-Binding Proteins - metabolism
HeLa Cells
Humans
Nuclear Proteins - metabolism
HeLa cells
DNA-PKcs
Ku70/80
Protein purification
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Summary: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.
ISSN:1940-6029
DOI:10.1007/978-1-0716-2063-2_14