In vitro characterisation of DNA binding of recombinant Rap2.4a to the redox box of the 2CPA promoter. (A) The 2CPA promoter region used in the Y1H-screen was amplified into 5 fragments by PCR (F1 – F5). Electrophoretic mobility shift assay (EMSA) was performed with 2.5 μg heterologously expressed His-tagged Rap2.4a or Rap2.6. The proteins were detected with anti-His antibody on Western-blots. (B) EMSA with a synthetic double-stranded oligonucleotide corresponding to 13 bp overlap of the fragments F4 and F5 (C) Similarity between AP-domain of Rap2.4a and other AP2-transcription factors according to PHYLIP. The maximal sequence variation is 22 %. (D) EMSA with the wild-type CE3-like element, its mutagenised variants MutA – MutE and an ABRE with His-tagged Rap2.4a followed by immunodetection with anti-His-antibody.