Sulfolobus acidocaldarius UDG Can Remove dU from the RNA Backbone: Insight into the Specific Recognition of Uracil Linked with Deoxyribose.

Gang-Shun Yi,Wei-Guo Cao,Xi-Peng Liu,Feng-Ping Wang,Wei-Wei Wang

doi:10.3390/genes8010038

Abstract

Sulfolobus acidocaldarius encodes family 4 and 5 uracil-DNA glycosylase (UDG). Two recombinant S. acidocaldarius UDGs (SacUDG) were prepared and biochemically characterized using oligonucleotides carrying a deaminated base. Both SacUDGs can remove deoxyuracil (dU) base from both double-stranded DNA and single-stranded DNA. Interestingly, they can remove U linked with deoxyribose from single-stranded RNA backbone, suggesting that the riboses on the backbone have less effect on the recognition of dU and hydrolysis of the C-N glycosidic bond. However, the removal of rU from DNA backbone is inefficient, suggesting strong steric hindrance comes from the 2′ hydroxyl of ribose linked to uracil. Both SacUDGs cannot remove 2,2′-anhydro uridine, hypoxanthine, and 7-deazaxanthine from single-stranded DNA and single-stranded DNA. Compared with the family 2 MUG, other family UDGs have an extra N-terminal structure consisting of about 50 residues. Removal of the 46 N-terminal residues of family 5 SacUDG resulted in only a 40% decrease in activity, indicating that the [4Fe-4S] cluster and truncated secondary structure are not the key elements in hydrolyzing the glycosidic bond. Combining our biochemical and structural results with those of other groups, we discussed the UDGs’ catalytic mechanism and the possible repair reactions of deaminated bases in prokaryotes.

Highlights

Some DNA damage in the genome is harmful to the cell if not repaired
U/G mismatch results from the hydrolysis deamination of the exocyclic amino group of cytosine in DNA, and a U/A base pair is generated via the misincorporation of dUMP into the DNA opposite base A during replication
Our results showed that two S. acidocaldarius uracil-DNA glycosylase (UDG) can remove dU residues from both single- and double-stranded DNA

Summary

Introduction

Some DNA damage in the genome is harmful to the cell if not repaired. Deoxyuracil (dU) is a kind of DNA damage and exists in the form of a U/G or U/A base pair. U/G mismatch results from the hydrolysis deamination of the exocyclic amino group of cytosine in DNA, and a U/A base pair is generated via the misincorporation of dUMP into the DNA opposite base A during replication. The G:U mismatched base pair will generate a permanent G:C to A:T transition mutation after replication. In addition to dU damage, hydrolysis deamination of the purines adenine and guanine inflicts damage on hypoxanthine and xanthine, respectively [1]. The rate constants for hydrolysis deamination of bases in DNA at elevated temperatures are several orders of magnitude higher than those at more moderate temperatures [2].

Results

Discussion

Conclusion