Analyses of protein bound to HRE-derivatized magnetic beads. (A) SDS-PAGE analysis. Nuclear extracts from hemin-treated U937 cells were incubated with magnetic beads derivatized with the 27-mer HRE DNA sequence as described in Materials and Methods. Bound proteins were released by boiling in Laemmli buffer, then separated by SDS-PAGE in 10% acrylamide gels. Protein bonds were visualized by silver or Coomassie blue staining. Lanes 1 and 2 depict results of two separate isolations. (B) Microsequence analysis of two peptides from the 90-kD HREBP band. Digests were sequenced on an Applied Biosystems 477A sequencer with an ABI 120 HPLC. KuAg p86 sequences are from the BLAST data base. Swiss-Prot accession:P13010; NCBI Seq ID:125731. (C) Immunoblot analyses of isolates in (A) with anti-KuAg monoclonal antibodies. Lane 1, Ab-3, clone 162 recognizing p70/p80; lane 2, Ab-4, clone N3H10 recognizing p70; lane 3, Ab-2, clone 11D recognizing p80.