Establishment of a cell-free system to study the activation of Chk2.
The checkpoint kinase Chk2 is activated in response to DNA damage through pathways requiring protein kinases ATM and/or ATR. The means by which Chk2 is activated by these kinases still remains to be addressed. Here we describe a cell-free system to study the activation of Chk2. Chk2 produced by a wheat germ extract in vitro transcription/translation system is inactive and can be activated by incubating with a rabbit reticulocyte lysate. This method will be useful for identification of cofactors required for activation of Chk2.
Xu
Xingzhi
X
Genetics of Development and Disease Branch, National Institute of Diabetes, Digestive and Kidney Diseases, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
Stern
David F
DF
D000818:Animals; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins; D002474:Cell-Free System; D064447:Checkpoint Kinase 2; D004249:DNA Damage; D004268:DNA-Binding Proteins; D005822:Genetic Vectors; D006801:Humans; D015151:Immunoblotting; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D014176:Protein Biosynthesis; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011817:Rabbits; D012156:Reticulocytes / Q000378:metabolism; D014158:Transcription, Genetic; D014908:Triticum / Q000378:metabolism; D025521:Tumor Suppressor Proteins
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D025521:Tumor Suppressor Proteins; D064447:Checkpoint Kinase 2; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C578634:CHEK2 protein, human; D017346:Protein Serine-Threonine Kinases