Negative superhelicity promotes ATP-dependent binding of yeast RAD3 protein to ultraviolet-damaged DNA

Patrick Sung, John F. Watkins, Louise Prakash, Satya Prakash

Research output: Contribution to journalArticle

28 Citations (Scopus)

Abstract

The RAD3 gene of Saccharomyces cerevisiae is required for excision repair of UV-damaged DNA and is essential for cell viability. Remarkable homology exists between RAD3 and the human excision repair gene XPD, whose mutational inactivation underlies the cancer-prone disorder in xeroderma pigmentosum group D patients. Our previous work demonstrated that RAD3-encoded protein contains a DNA helicase activity. Here, we show that RAD3 binds preferentially to UV-damaged DNA over nondamaged DNA. Removal of pyrimidine dimers from damaged DNA by enzymatic photoreactivation does not affect binding, suggesting an affinity of RAD3 for pyrimidine (6-4) pyrimidone photoproducts. Damage-specific binding by RAD3 is strongly dependent on ATP and on the degree of negative superhelicity in DNA. The requirement of superhelicity in damage binding may target RAD3 to regions of DNA undergoing transcription, resulting in the preferential repair of these regions. The rad3 Arg-48 mutant protein, which lacks the DNA helicase activity, also binds UV-damaged DNA preferentially, indicating that DNA helicase and damage binding are two distinct and separable functional entities in RAD3.

Original languageEnglish (US)
Pages (from-to)8303-8308
Number of pages6
JournalJournal of Biological Chemistry
Volume269
Issue number11
StatePublished - Mar 18 1994

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Fungal Proteins
Adenosine Triphosphate
DNA Helicases
DNA
Repair
DNA Repair
Genes
Pyrimidinones
Pyrimidine Dimers
Xeroderma Pigmentosum
Mutant Proteins
Transcription
Yeast
DNA Damage
Saccharomyces cerevisiae
Cell Survival
Cells
Neoplasms

ASJC Scopus subject areas

  • Biochemistry

Cite this

Negative superhelicity promotes ATP-dependent binding of yeast RAD3 protein to ultraviolet-damaged DNA. / Sung, Patrick; Watkins, John F.; Prakash, Louise; Prakash, Satya.

In: Journal of Biological Chemistry, Vol. 269, No. 11, 18.03.1994, p. 8303-8308.

Research output: Contribution to journalArticle

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