Characterization of reaction intermediates of human excision repair nuclease.

Published

Journal Article

Nucleotide excision repair in humans is a complex reaction involving 14 polypeptides in six repair factors for dual incisions on either sides of a DNA lesion. To identify the reaction intermediates that form by the human excision repair nuclease, we adopted three approaches: purification of functional DNA.protein complexes, permanganate footprinting, and the employment as substrate of presumptive DNA reaction intermediates containing unwound sequences 5' to, 3' to, or encompassing the DNA lesion. The first detectable reaction intermediate was formed by substrate binding of XPA, RPA, XPC.HHR23B plus TFIIH (preincision complex 1, PIC1). In this complex the DNA was unwound on either side of the lesion by no more than 10 bases. Independent of the XPG nuclease function, the XPG protein stabilized this complex, forming a long lived preincision complex 2 (PIC2). The XPF.ERCC1 complex bound to PIC2, forming PIC3, which led to dual incisions and the release of the excised oligomer. With partially unwound DNAs, thymine cyclobutane dimer was excised at a fast rate independent of XPC.HHR23B, indicating that a major function of this protein is to stabilize the unwound DNA or to aid lesion unwinding in preincision complexes.

Full Text

Duke Authors

Cited Authors

  • Mu, D; Wakasugi, M; Hsu, DS; Sancar, A

Published Date

  • November 14, 1997

Published In

Volume / Issue

  • 272 / 46

Start / End Page

  • 28971 - 28979

PubMed ID

  • 9360969

Pubmed Central ID

  • 9360969

International Standard Serial Number (ISSN)

  • 0021-9258

Digital Object Identifier (DOI)

  • 10.1074/jbc.272.46.28971

Language

  • eng

Conference Location

  • United States