A novel four zinc-finger protein targeted against p190(BcrAbl) fusion oncogene cDNA: utilisation of zinc-finger recognition codes.
A three zinc-finger protein that binds specifically to the cDNA representing the unique fusion gene BCR:Abl, associated with acute lymphoblastic leukaemia, has previously been characterised. At this breakpoint, a sequence homology of 8/9 bp exists between the BCR:Abl (fusion) and c-ABL: (parental) target sequences. We show that the three zinc-finger protein discriminates poorly between the fusion (BCR:Abl) and parental (ABL:) sequence (K:(d)s of 42.8 and 65.1 nM, respectively). In order to improve the discriminatory properties of this protein, and to demonstrate the utility of current zinc-finger databases, we have added a fourth zinc-finger to the original three zinc-finger protein. This fourth finger recognises a 3 bp subsite derived from the BCR: portion of the breakpoint and is not present in c-ABL: This novel four finger protein, which now recognises a 12 bp sequence, demonstrates improved specific binding to BcrAbl (K:(d )= 17 nM). More significantly we have shown that there is now enhanced discrimination between BcrAbl and ABL: sequences by the four finger protein than the original three finger protein.
McNamara
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Department of Molecular Medicine, The Rayne Institute, Guy's, King's and St Thomas' Schools of Medicine and Dentistry, 123 Coldharbour Lane, Camberwell, London SE5 9NU, UK.
Ford
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D000595:Amino Acid Sequence; D001483:Base Sequence; D001667:Binding, Competitive; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016044:Fusion Proteins, bcr-abl / Q000235:genetics*; D016313:Genes, abl / Q000235:genetics; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D015513:Oncogene Proteins / Q000235:genetics; D009857:Oncogenes / Q000235:genetics*; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases; D011518:Proto-Oncogene Proteins; D051562:Proto-Oncogene Proteins c-bcr; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D013816:Thermodynamics; D016335:Zinc Fingers
D004268:DNA-Binding Proteins; D015513:Oncogene Proteins; D011518:Proto-Oncogene Proteins; D011993:Recombinant Fusion Proteins; D004247:DNA; D011505:Protein-Tyrosine Kinases; D016044:Fusion Proteins, bcr-abl; C496613:BCR protein, human; D051562:Proto-Oncogene Proteins c-bcr