A novel matrix attachment region DNA binding motif identified using a random phage peptide library

Abstract

SATB1 is a nuclear matrix attachment DNA (MAR). binding protein which is predominantly expressed in thymocytes. This protein binds to the minor groove specifically recognizing an unusual DNA context exhibited by a specific MAR region with strong base-unpairing propensity. A phage library displaying nonamer random peptides without any built-in structure was used to identify a MAR binding motif of SATB1. One predominant cyclic peptide C1 of CRQNWGLEGC selected by a MAR-affinity column showed 50% identity with a segment in SATB1 (amino acids 355-363). Replacement of the C1 similarity segment in SATB1 by a random amino acid sequence or its truncation resulted in more than 80% reduction in MAR binding. In contrast, replacement of the same SATB1 segment with the C1 peptide restored full MAR binding activity and specificity as the wildtype protein. Single amino acid mutation of the conserved Arg or Glu residue to Ala greatly reduced MAR binding. Taken together our data show that a nine amino acid sequence in SATB1 represents a key MAR binding motif. Phage display may provide a general tool for rapid identification of DNA binding peptide motifs.