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PMID:16595800

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Citation

Masuda-Sasa, T, Imamura, O and Campbell, JL (2006) Biochemical analysis of human Dna2. Nucleic Acids Res. 34:1865-75

Abstract

Yeast Dna2 helicase/nuclease is essential for DNA replication and assists FEN1 nuclease in processing a subset of Okazaki fragments that have long single-stranded 5' flaps. It is also involved in the maintenance of telomeres. DNA2 is a gene conserved in eukaryotes, and a putative human ortholog of yeast DNA2 (ScDNA2) has been identified. Little is known about the role of human DNA2 (hDNA2), although complementation experiments have shown that it can function in yeast to replace ScDNA2. We have now characterized the biochemical properties of hDna2. Recombinant hDna2 has single-stranded DNA-dependent ATPase and DNA helicase activity. It also has 5'-3' nuclease activity with preference for single-stranded 5' flaps adjacent to a duplex DNA region. The nuclease activity is stimulated by RPA and suppressed by steric hindrance at the 5' end. Moreover, hDna2 shows strong 3'-5' nuclease activity. This activity cleaves single-stranded DNA in a fork structure and, like the 5'-3' activity, is suppressed by steric hindrance at the 3'-end, suggesting that the 3'-5' nuclease requires a 3' single-stranded end for activation. These biochemical specificities are very similar to those of the ScDna2 protein, but suggest that the 3'-5' nuclease activity may be more important than previously thought.

Links

PubMed PMC1428797 Online version:10.1093/nar/gkl070

Keywords

Adenosine Triphosphatases/genetics; Adenosine Triphosphatases/isolation & purification; Adenosine Triphosphatases/metabolism; Animals; Baculoviridae/genetics; Cell Line; DNA Helicases/genetics; DNA Helicases/isolation & purification; DNA Helicases/metabolism; DNA, Single-Stranded/metabolism; Deoxyribonucleases/genetics; Deoxyribonucleases/isolation & purification; Deoxyribonucleases/metabolism; Endodeoxyribonucleases/genetics; Endodeoxyribonucleases/isolation & purification; Endodeoxyribonucleases/metabolism; Exodeoxyribonucleases/genetics; Exodeoxyribonucleases/isolation & purification; Exodeoxyribonucleases/metabolism; HeLa Cells; Humans; Insects/cytology; Recombinant Proteins/isolation & purification; Recombinant Proteins/metabolism; Substrate Specificity

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

HUMAN:DNA2

enables

GO:0017116: single-stranded DNA helicase activity

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

HUMAN:DNA2

enables

GO:0003678: DNA helicase activity

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

HUMAN:DNA2

enables

GO:0017108: 5'-flap endonuclease activity

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

HUMAN:DNA2

enables

GO:0043142: single-stranded DNA-dependent ATPase activity

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

Notes

See also

References

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