RPA and XPA interaction with DNA structures mimicking intermediates of the late stages in nucleotide excision repair

Yuliya S. Krasikova, Nadejda I. Rechkunova, Ekaterina A. Maltseva, Olga I. Lavrik

Research output: Contribution to journalArticlepeer-review

6 Citations (Scopus)

Abstract

Replication protein A (RPA) and the xeroderma pigmentosum group A (XPA) protein are indispensable for both pathways of nucleotide excision repair (NER). Here we analyze the interaction of RPA and XPA with DNA containing a flap and different size gaps that imitate intermediates of the late NER stages. Using gel mobility shift assays, we found that RPA affinity for DNA decreased when DNA contained both extended gap and similar sized flap in comparison with gapped-DNA structure. Moreover, crosslinking experiments with the flap-gap DNA revealed that RPA interacts mainly with the ssDNA platform within the long gap and contacts flap in DNA with a short gap. XPA exhibits higher affinity for bubble-DNA structures than to flap-gap-containing DNA. Protein titration analysis showed that formation of the RPA-XPA-DNA ternary complex depends on the protein concentration ratio and these proteins can function as independent players or in tandem. Using fluorescently-labelled RPA, direct interaction of this protein with XPA was detected and characterized quantitatively. The data obtained allow us to suggest that XPA can be involved in the post-incision NER stages via its interaction with RPA.

Original languageEnglish
Article number0190782
Pages (from-to)e0190782
Number of pages20
JournalPLoS ONE
Volume13
Issue number1
DOIs
Publication statusPublished - 10 Jan 2018

Keywords

  • DNA Repair
  • DNA/chemistry
  • Electrophoretic Mobility Shift Assay
  • Humans
  • Photoaffinity Labels
  • Protein Binding
  • Recombinant Proteins/metabolism
  • Replication Protein A/metabolism
  • Xeroderma Pigmentosum Group A Protein/metabolism
  • DAMAGED DNA
  • COMPLEX
  • RECOGNITION
  • REPLICATION PROTEIN-A
  • SINGLE-STRANDED-DNA
  • BINDING PROTEIN
  • OLIGONUCLEOTIDES
  • DUAL INCISION
  • TFIIH
  • MOLECULAR-MECHANISMS

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