Fig. 5

Electrophoretic mobility shift assay of AP-1. The DNA-binding activity of AP-1 was estimated using an electrophoretic mobility shift assay. NB9 was incubated with 50 µM or 2 mM L-Arg for 48 hr. DNA extracts from each of these cells were incubated with an AP-1-specific ³²P-oligonucleotide for 20 min and then loaded on a 6% nondenaturing Polyacrylamide gel and subjected to electrophoresis in 25 mM Tris, 22.5 mM borate, and 0.25 mM EDTA, pH 8.0. The DNA binding activity of the extracts was quantified by estimating the amount of the ³²P-labeled AP-1. Lane 1, free probe; lane 2, Hela cell extracts as a positive control; lane 3, a 100-fold excess of unlabeled probe as a specificity control; lane 4, 50 µM L-Arg; lane 5, 2 mM L-Arg.