Taylor, John S. and Schultz, Peter G. and Dervan, Peter B. (1984) DNA affinity cleaving : Sequence specific cleavage of DNA by Distamycin-EDTA • Fe(II) and EDTA-distamycin • Fe(II). Tetrahedron, 40 (3). pp. 457-465. ISSN 0040-4020. doi:10.1016/0040-4020(84)85050-4. https://resolver.caltech.edu/CaltechAUTHORS:20160511-152456776
Full text is not posted in this repository. Consult Related URLs below.
Use this Persistent URL to link to this item: https://resolver.caltech.edu/CaltechAUTHORS:20160511-152456776
Abstract
The attachment of EDTA· Fe(II) to distamycin changes the sequence specific DNA binding antibiotic into a sequence specific DNA cleaving molecule. We report the synthesis of EDTA-distamycin (ED) which has the metal chelator, EDTA, tethered to the carboxy terminus of the N-methylpyrrole tripeptide moiety of the antiobiotic, distamycin. EDTA-distamycin • Fe(II) (ED · Fe II) at 10^(-6) M concentration efficiently cleaves pBR322 DNA (10^(-5) M in base pairs) in the presence of oxygen and dithiothreitol (DTT). Using Maxam-Gilbert sequencing gel analyses, we find that ED · Fe (II) affords DNA cleavage patterns of unequal intensity covering two to four contiguous base pairs adjacent to a five base pair site consisting of adenines (A) and thymines (T). The multiple cleavages at each site might be evidence for a diffusible oxidizing species, perhaps hydroxyl radical. The unequal intensity of cleavage on each side of the A + T site permit assignment of major and minor orientations of the tripeptide binding unit. A comparison of the cleavage specificity of ED · Fe (II) with distamycin-EDTA · Fe (II), (DE · Fe (II)) which has EDTA · Fe (II) attached to the amino terminus of the N-methylpyrrole tripeptide, shows DNA cleavage patterns at the same sites but with intensities of opposite polarity. Maxam-Gilbert sequencing gel analysis of the DNA cleavage patterns by ED • Fe (II) and DE • Fe (II) on both DNA strands of a 381 base pair restriction fragment reveals asymmetric DNA cleavage patterns. Cleavage is shifted to the 3' side of each DNA strand. A model consistent with this cleavage pattern indicates one preferred binding te for ED • Fe (II) and DE • Fe (II) is 3'-TTTAA-5' with the “amino end” of the tripeptide oriented to the 3' end of the thymine rich strand. This “DNA affinity cleavage” method which consists of attaching cleaving functions to DNA binding molecules followed by DNA cleavage pattern analyses using Maxam-Gilbert sequencing gels may be a useful direct method for determining the binding site and orientation of small molecules on native DNA.
Item Type: | Article | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Related URLs: |
| |||||||||
ORCID: |
| |||||||||
Additional Information: | © 1984 Elsevier Ltd. Received 1 October 1982. Supported by Fellowship DRG-526 of the Damon Runyon-Walter Winchell Cancer Fund. | |||||||||
Funders: |
| |||||||||
Issue or Number: | 3 | |||||||||
DOI: | 10.1016/0040-4020(84)85050-4 | |||||||||
Record Number: | CaltechAUTHORS:20160511-152456776 | |||||||||
Persistent URL: | https://resolver.caltech.edu/CaltechAUTHORS:20160511-152456776 | |||||||||
Official Citation: | John S. Taylor, Peter G. Schultz, Peter B. Dervan, DNA affinity cleaving , Tetrahedron, Volume 40, Issue 3, 1984, Pages 457-465, ISSN 0040-4020, http://dx.doi.org/10.1016/0040-4020(84)85050-4. (http://www.sciencedirect.com/science/article/pii/0040402084850504) | |||||||||
Usage Policy: | No commercial reproduction, distribution, display or performance rights in this work are provided. | |||||||||
ID Code: | 67023 | |||||||||
Collection: | CaltechAUTHORS | |||||||||
Deposited By: | Victoria Brennan | |||||||||
Deposited On: | 19 May 2016 19:16 | |||||||||
Last Modified: | 11 Nov 2021 00:25 |
Repository Staff Only: item control page