Substrate specificity of the Rad3 ATPase/DNA helicase of Saccharomyces cerevisiae and binding of Rad3 protein to nucleic acids

Hanspeter Naegeli, Lee Bardwell, Itzik Harosh, Errol C. Friedberg

Research output: Contribution to journalArticle

15 Citations (Scopus)

Abstract

Rad3 protein from the yeast Saccharomyces cerevisiae is a single-stranded DNA-dependent ATPase which catalyzes the unwinding of DNA-DNA duplexes. In the present studies we have demonstrated that the purified enzyme additionally catalyzes the displacement of RNA fragments annealed to complementary DNA. Quantitative comparisons using otherwise identical partially duplex DNA-DNA and DNA-RNA substrates indicate a significant preference for the latter. Competition for ATPase or DNA helicase activity by various homopolymers suggests that Rad3 protein does not discriminate between ribonucleotide and deoxyribonucleotide homopolymers with respect to binding. However, neither single-stranded RNA nor various ribonucleotide homopolymers supported the hydrolysis of nucleoside 5′-triphosphates. Additionally, Rad3 protein was unable to catalyze the displacement of oligo(dA) annealed to poly(U), suggesting that the catalytic domain of the enzyme is exquisitely sensitive to chemical and/or or conformational differences between DNA and RNA. Hence, it appears that Rad3 protein is not an RNA helicase.

Original languageEnglish (US)
Pages (from-to)7839-7844
Number of pages6
JournalJournal of Biological Chemistry
Volume267
Issue number11
StatePublished - Apr 15 1992

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DNA Helicases
Substrate Specificity
Yeast
Nucleic Acids
Saccharomyces cerevisiae
Adenosine Triphosphatases
Carrier Proteins
DNA
Substrates
Homopolymerization
RNA
Ribonucleotides
Proteins
Deoxyribonucleotides
RNA Helicases
Saccharomyces cerevisiae Proteins
Poly U
Fungal Proteins
Enzymes
Nucleosides

ASJC Scopus subject areas

  • Biochemistry

Cite this

Substrate specificity of the Rad3 ATPase/DNA helicase of Saccharomyces cerevisiae and binding of Rad3 protein to nucleic acids. / Naegeli, Hanspeter; Bardwell, Lee; Harosh, Itzik; Friedberg, Errol C.

In: Journal of Biological Chemistry, Vol. 267, No. 11, 15.04.1992, p. 7839-7844.

Research output: Contribution to journalArticle

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N2 - Rad3 protein from the yeast Saccharomyces cerevisiae is a single-stranded DNA-dependent ATPase which catalyzes the unwinding of DNA-DNA duplexes. In the present studies we have demonstrated that the purified enzyme additionally catalyzes the displacement of RNA fragments annealed to complementary DNA. Quantitative comparisons using otherwise identical partially duplex DNA-DNA and DNA-RNA substrates indicate a significant preference for the latter. Competition for ATPase or DNA helicase activity by various homopolymers suggests that Rad3 protein does not discriminate between ribonucleotide and deoxyribonucleotide homopolymers with respect to binding. However, neither single-stranded RNA nor various ribonucleotide homopolymers supported the hydrolysis of nucleoside 5′-triphosphates. Additionally, Rad3 protein was unable to catalyze the displacement of oligo(dA) annealed to poly(U), suggesting that the catalytic domain of the enzyme is exquisitely sensitive to chemical and/or or conformational differences between DNA and RNA. Hence, it appears that Rad3 protein is not an RNA helicase.

AB - Rad3 protein from the yeast Saccharomyces cerevisiae is a single-stranded DNA-dependent ATPase which catalyzes the unwinding of DNA-DNA duplexes. In the present studies we have demonstrated that the purified enzyme additionally catalyzes the displacement of RNA fragments annealed to complementary DNA. Quantitative comparisons using otherwise identical partially duplex DNA-DNA and DNA-RNA substrates indicate a significant preference for the latter. Competition for ATPase or DNA helicase activity by various homopolymers suggests that Rad3 protein does not discriminate between ribonucleotide and deoxyribonucleotide homopolymers with respect to binding. However, neither single-stranded RNA nor various ribonucleotide homopolymers supported the hydrolysis of nucleoside 5′-triphosphates. Additionally, Rad3 protein was unable to catalyze the displacement of oligo(dA) annealed to poly(U), suggesting that the catalytic domain of the enzyme is exquisitely sensitive to chemical and/or or conformational differences between DNA and RNA. Hence, it appears that Rad3 protein is not an RNA helicase.

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