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Published May 1994 | metadata_only
Journal Article

Synthesis of a hybrid protein containing the iron-binding ligand of bleomycin and the DNA-binding domain of Hin


The iron-binding and oxygen-activating domain of the natural product bleomycin [pyrimidoblamic acid-beta-hydroxy-L-histidine (PBA-beta-OH-His)] was attached to the NH_2 terminus of the DNA binding domain of Hin recombinase (residues 139-190). This hybrid 54-residue protein PBA-beta-OH-His-Hin-(139-190) binds specifically to DNA at four distinct Hin binding sites with affinities comparable to those of the unmodified Hin(139-190). In the presence of dithiothreitol, Fe(II).PBA-beta-OH-His-Hin-(139-190) cleaves DNA with specificity remarkably similar to that of Fe(II).EDTA-Hin(139-190). Analysis of the cleavage patterns suggests that site-specific DNA cleavage is mediated by a localized diffusible species, in contrast with cleavage by bleomycin, which occurs through a nondiffusible oxidant. This has implications for the design of second-generation artificial sequence specific DNA cleaving proteins and defines limitations in current efforts to create atom-specific chemistry on DNA.

Additional Information

© 1994 American Chemical Society. Received February 16, 1994. Abstract published in Advance ACS Abstracts, April 15, 1994. This work was supported by the National Institutes of Health (GM-276811, a National Science Foundation predoctoral fellowship, a National Institutes of Health predoctoral traineeship, and a Glaxo Graduate Student Fellowship to M.G.O. and an American Cancer Society postdoctoral fellowship to K.D.T. We thank G. M. Hathaway (University of California, Riverside) for protein mass spectral analysis.

Additional details

August 20, 2023
August 20, 2023