title: The designed protein M(II)-Gly-Lys-His-Fos(138-211) specifically cleaves the AP-1 binding site containing DNA.
abstract: A new specific DNA cleavage protein, Gly-Lys-His-Fos(138-211), was designed, expressed, and characterized. The DNA-binding component of the design uses the basic and leucine zipper regions of the leucine zipper Fos, which are represented by Fos(138-211). The DNA cleavage moiety was provided by the design of the amino-terminal Cu(II)-, Ni(II)-binding site GKH at the amino terminus of Fos(138-211). Binding of Cu(II) or Ni(II) by the protein activates its cleavage ability. The GKH motif was predicted to form a specific amino-terminal Cu(II)-, Ni(II)-binding motif as previously defined [Predki, P. F., Harford, C., Brar, P., & Sarkar, B. (1992) Biochem. J. 287, 211 -215]. This prediction was verified as the tripeptide, GKH, and the expressed protein, GKH-Fos(138-211), were both shown to be capable of binding Cu(II) and Ni(II). The designed protein upon heterodimerization with Jun(248-334) was shown to bind to and cleave several forms of DNA which contained an AP-1 binding site. The cleavage was shown to be specific. This design demonstrates the versatility of the amino-terminal Cu(II)-, Ni(II)-binding motif and the variety of motifs which can be generated. The site of cleavage by GKH-Fos(138-211) on DNA provides further information regarding the bending of DNA upon binding to Fos-Jun heterodimers.
authors.affiliation: Department of Biochemistry Research, The Hospital for Sick Children, Toronto, Ontario, Canada.
authors.forename: C
authors.initials: C
authors.lastname: Harford
authors.forename: S
authors.initials: S
authors.lastname: Narindrasorasak
authors.forename: B
authors.initials: B
authors.lastname: Sarkar
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015195:Drug Design; D066298:In Vitro Techniques; D016350:Leucine Zippers / Q000235:genetics; D008670:Metals / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016760:Proto-Oncogene Proteins c-fos / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016755:Proto-Oncogene Proteins c-jun / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018808:Transcription Factor AP-1 / Q000378:metabolism*
chemical_list: D008670:Metals; D011506:Proteins; D016760:Proto-Oncogene Proteins c-fos; D016755:Proto-Oncogene Proteins c-jun; D018808:Transcription Factor AP-1; D004247:DNA