Synthesis of a hybrid protein containing the iron-binding ligand of bleomycin and the DNA-binding domain of Hin
Abstract
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
- Eprint ID
- 66919
- DOI
- 10.1021/bc00027a009
- Resolver ID
- CaltechAUTHORS:20160510-133747674
- GM-276811
- NIH
- NSF Predoctoral Fellowship
- NIH Predoctoral Fellowship
- Glaxo
- American Cancer Society
- Created
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2016-05-18Created from EPrint's datestamp field
- Updated
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2021-11-11Created from EPrint's last_modified field