Skip NavigationSkip to Content
Return Return to Search Results

The DNA binding and 3'-end preferential activity of human tyrosyl-DNA phosphodiesterase

  1. Author:
    Dexheimer, T. S.
    Stephen, A. G.
    Fivash, M. J.
    Fisher, R. J.
    Pommier, Y.

  2. Author Address

    [Dexheimer, Thomas S.; Pommier, Yves] NCI, Mol Pharmacol Lab, Ctr Canc Res, NIH, Bethesda, MD 20892 USA. [Stephen, Andrew G.; Fisher, Robert J.] SAIC Frederick Inc, Prot Chem Lab, Adv Technol Program, Frederick, MD 21702 USA. [Fivash, Matthew J.] Data Management Syst Inc, NCI Frederick, Frederick, MD 21702 USA.;Pommier, Y, NCI, Mol Pharmacol Lab, Ctr Canc Res, NIH, 37 Convent Dr,Bldg 37,Room 5068, Bethesda, MD 20892 USA.;pommier@nih.gov
    1. Year: 2010
    2. Date: Apr
  2. Journal: Nucleic Acids Research
    1. 38
    2. 7
    3. Pages: 2444-2452
  4. Type of Article: Article
  5. ISSN: 0305-1048
  1. Abstract:

    Human tyrosyl-DNA phosphodiesterase (Tdp1) processes 3'-blocking lesions, predominantly 3'-phosphotyrosyl bonds resulting from the trapping of topoisomerase I (Top1) cleavage complexes. The controversial ability of yeast Tdp1 to hydrolyze 5'-phosphotyrosyl linkage between topoisomerase II (Top2) and DNA raises the question whether human Tdp1 possesses 5'-end processing activity. Here we characterize the end-binding and cleavage preference of human Tdp1 using single-stranded 5'- and 3'-fluorescein-labeled oligonucleotides. We establish 3'-fluorescein as an efficient surrogate substrate for human Tdp1, provided it is attached to the DNA by a phosphodiester (but not a phosphorothioate) linkage. We demonstrate that human Tdp1 lacks the ability to hydrolyze a phosphodiester linked 5'-fluorescein. Using both fluorescence anisotropy and time-resolved fluorescence quenching techniques, we also show the preferential binding of human Tdp1 to the 3'-end. However, DNA binding competition experiments indicate that human Tdp1 binding is dependent on DNA length rather than number of DNA ends. Lastly, using surface plasmon resonance, we show that human Tdp1 selectively binds the 3'-end of DNA. Together, our results suggest human Tdp1 may act using a scanning mechanism, in which Tdp1 bind non-specifically upstream of a 3'-blocking lesion and is preferentially stabilized at 3'-DNA ends corresponding to its site of action.

    See More

  1. Keywords:

External Sources

  1. View Full Text
  2. DOI: 10.1093/nar/gkp1206
  3. View record in Web of Science®
  4. WOS: 000276744600038

Library Notes

  1. Fiscal Year: FY2009-2010
NCI at Frederick

You are leaving a government website.

This external link provides additional information that is consistent with the intended purpose of this site. The government cannot attest to the accuracy of a non-federal site.

Linking to a non-federal site does not constitute an endorsement by this institution or any of its employees of the sponsors or the information and products presented on the site. You will be subject to the destination site's privacy policy when you follow the link.

ContinueCancel