Mack, David P. and Dervan, Peter B. (1992) Sequence-specific oxidative cleavage of DNA by a designed metalloprotein, nickel(II)•GGH(Hin139-190). Biochemistry, 31 (39). pp. 9399-405. ISSN 0006-2960. doi:10.1021/bi00154a011. https://resolver.caltech.edu/CaltechAUTHORS:20160510-142115186
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Abstract
A 55-residue protein containing the DNA binding domain of Hin recombinase, residues 139-190, with the tripeptide Gly-Gly-His (GGH) at the NH_2 terminus was synthesized by stepwise solid-phase methods. GGH(Hin139-190) binds sequence specifically to DNA at four 13 base pair sites (termed hixL and secondary) and, in the presence of Ni(OAc)_2 and monoperoxyphthalic acid, reacts predominantly at a single deoxyribose position on one strand of each binding site [Mack, D.P., & Dervan, P.B. (1990) J. Am. Chem. Soc. 112, 4604]. We find that, upon treatment with n-butylamine, the DNA termini at the cleavage site are 3'- and 5'-phosphate, consistent with oxidative degradation of the deoxyribose backbone. The nickel-mediated oxidation can be activated with peracid, iodosylbenzene, or hydrogen peroxide. The sequence specificity of the reaction is not dependent on oxidant, but the rates of cleavage differ, decreasing in the order peracid greater than iodosylbenzene greater than hydrogen peroxide. Optimal cleavage conditions for a 1 µM concentration of protein are 50 µM peracid, pH 8.0, and 1 equiv of Ni(OAc)_2. The preferential cleavage at a single base pair position on one strand of the minor groove indicates a nondiffusible oxidizing species. A change of absolute configuration in the GGH metal binding domain from L-His to D-His [Ni(II)•GG-(-D-)H(Hin139-190)] affords cleavage at similar base pair locations but opposite with regard to strand specificity.
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Additional Information: | 0 1992 American Chemical Society. Received May 26, 1992. We are grateful to the National Institutes of Health (GM27681) and the National Foundation for Cancer Research for financial support. | |||||||||
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Issue or Number: | 39 | |||||||||
DOI: | 10.1021/bi00154a011 | |||||||||
Record Number: | CaltechAUTHORS:20160510-142115186 | |||||||||
Persistent URL: | https://resolver.caltech.edu/CaltechAUTHORS:20160510-142115186 | |||||||||
Official Citation: | Sequence-specific oxidative cleavage of DNA by a designed metalloprotein, nickel(II).cntdot.GGH(Hin139-190) David P. Mack and Peter B. Dervan Biochemistry 1992 31 (39), 9399-9405 DOI: 10.1021/bi00154a011 | |||||||||
Usage Policy: | No commercial reproduction, distribution, display or performance rights in this work are provided. | |||||||||
ID Code: | 66942 | |||||||||
Collection: | CaltechAUTHORS | |||||||||
Deposited By: | Victoria Brennan | |||||||||
Deposited On: | 19 May 2016 22:46 | |||||||||
Last Modified: | 11 Nov 2021 00:24 |
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