JNK Kinase Assay

THE BASIC PROTOCOL IS THE SAME AS THE MAP KINASE ASSAY, with the following changes:

REACTION BUFFER

25mM HEPES pH 7.4

25mM b-glycerophosphate (not needed)

50nM OA

25mM MgCl2

0.1mM Na3VO4

0.5mM DTT

Kinase assay: 10ml beads

26ml Reactionbuffer

2ml GST-Jun (5mg/ml)

1ml 1mM ATP

1ml [g³²P]-ATP (10mCi) (5mCi is OK)

Incubate 15 minutes at Room temp

Stop with SDS sample buffer

SDS PAGE and autoradiography