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XRCC1 stimulates polynucleotide kinase by enhancing its damage discrimination and displacement from DNA repair intermediates

journal contribution
posted on 2023-06-07, 21:00 authored by Rajam S Mani, Mesfin Fanta, Feridoun Karimi-Busheri, Elizabeth Silver, César A Virgen, Keith CaldecottKeith Caldecott, Carol E Cass, Michael Weinfeld
Human polynucleotide kinase (hPNK) is required for processing and rejoining DNA strand break termini. The 5'-DNA kinase and 3'-phosphatase activities of hPNK can be stimulated by the ¿scaffold¿ protein XRCC1, but the mechanism remains to be fully elucidated. Using a variety of fluorescence techniques, we examined the interaction of hPNK with XRCC1 and substrates that model DNA single-strand breaks. hPNK binding to substrates with 5'-OH termini was only ~5-fold tighter than that to identical DNA molecules with 5'-phosphate termini, suggesting that hPNK remains bound to the product of its enzymatic activity. The presence of XRCC1 did not influence the binding of hPNK to substrates with 5'-OH termini, but sharply reduced the interaction of hPNK with DNA bearing a 5'-phosphate terminus. These data, together with kinetic data obtained at limiting enzyme concentration, indicate a dual function for the interaction of XRCC1 with hPNK. First, XRCC1 enhances the capacity of hPNK to discriminate between strand breaks with 5'-OH termini and those with 5'-phosphate termini; and second, XRCC1 stimulates hPNK activity by displacing hPNK from the phosphorylated DNA product.

History

Publication status

  • Published

Journal

Journal of Biological Chemistry

ISSN

0021-9258

Issue

38

Volume

282

Page range

28004-28013

Pages

10.0

Department affiliated with

  • Sussex Centre for Genome Damage Stability Publications

Full text available

  • No

Peer reviewed?

  • Yes

Legacy Posted Date

2012-02-06

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