Dataset Information

Uracil DNA glycosylase uses DNA hopping and short-range sliding to trap extrahelical uracils.

ABSTRACT: The astonishingly efficient location and excision of damaged DNA bases by DNA repair glycosylases is an especially intriguing problem in biology. One example is the enzyme uracil DNA glycosylase (UNG), which captures and excises rare extrahelical uracil bases that have emerged from the DNA base stack by spontaneous base pair breathing motions. Here, we explore the efficiency and mechanism by which UNG executes intramolecular transfer and excision of two uracil sites embedded on the same or opposite DNA strands at increasing site spacings. The efficiency of intramolecular site transfer decreased from 41 to 0% as the base pair spacing between uracil sites on the same DNA strand increased from 20 to 800 bp. The mechanism of transfer is dominated by DNA hopping between landing sites of approximately 10 bp size, over which rapid 1D scanning likely occurs. Consistent with DNA hopping, site transfer at 20- and 56-bp spacings was unaffected by whether the uracils were placed on the same or opposite strands. Thus, UNG uses hopping and 3D diffusion through bulk solution as the principal pathways for efficient patrolling of long genomic DNA sequences for damage. Short-range sliding over the range of a helical turn allows for redundant inspection of very local DNA sequences and trapping of spontaneously emerging extrahelical uracils.

SUBMITTER: Porecha RH

PROVIDER: S-EPMC2504779 | biostudies-literature | 2008 Aug

REPOSITORIES: biostudies-literature

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Uracil DNA glycosylase uses DNA hopping and short-range sliding to trap extrahelical uracils.

Porecha Rishi H RH Stivers James T JT

Proceedings of the National Academy of Sciences of the United States of America 20080731 31

The astonishingly efficient location and excision of damaged DNA bases by DNA repair glycosylases is an especially intriguing problem in biology. One example is the enzyme uracil DNA glycosylase (UNG), which captures and excises rare extrahelical uracil bases that have emerged from the DNA base stack by spontaneous base pair breathing motions. Here, we explore the efficiency and mechanism by which UNG executes intramolecular transfer and excision of two uracil sites embedded on the same or oppos ...[more]

PMID: 18669665

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Project description:Two dimensional (2D) NMR and molecular dynamics simulations have been used to determine the three dimensional (3D) structure of a hairpin DNA, d-CTA-GAGGATCC-TUTT-GGATCCT (22mer; abbreviated as U2-hairpin), which has uracil at the second position from the 5' end of the tetraloop. The(1)H resonances of this hairpin have been assigned almost completely. NMR restrained molecular dynamics and energy minimization procedures have been used to describe the 3D structure of U2-hairpin. This study establishes that the stem of the hairpin adopts a right-handed B-DNA conformation, while the T(12)and T(15)nucleotides stack upon 3' and 5' ends of the stem, respectively. Further, T(14)stacks upon both T(12)and T(15). Though U(13)partially stacks upon T(14), no stacking interaction is observed between U(13)and T(12). All the individual nucleotide bases belonging to the stem and T(12)and T(15)of the loop adopt ' anti ' conformation with respect to their sugar moiety, while the U(13)and T(14)of the loop are in ' syn ' conformation. The turning phosphate in the loop is located between T(13)and T(14). This study and a concurrent NMR structural study on yet another hairpin DNA d-CTAGAGGAATAA-TTTU-GGATCCT (22mer; abbreviated as U4-hairpin), with uracil at the fourth position from the 5' end of the tetraloop throw light upon various interactions which have been reported between Escherichia coli uracil DNA glycosylase (UDG) and uracil containing DNA. The epsilon of T(12)and alpha, beta, gamma, epsilon and zeta of U(13)and gamma of T(14), which partially influence the local conformation of U(13)in U2-hairpin are all locked in ' trans ' conformation. Such stretched out backbone conformation in the vicinity of U(13)could be the reason as to why the U2-hairpin is found to be the poor substrate for its interaction with UDG compared to the other substrates in which the uracil is at first, third and fourth positions of the tetraloop from its 5' end, as reported earlier by Vinay and Varshney. This study shows that UDG actively promotes the flipping of uracil from a stacked conformation and rules out the possibility of UDG recognizing the flipped out uracil bases.

Dataset Information

Uracil DNA glycosylase uses DNA hopping and short-range sliding to trap extrahelical uracils.

Publications

Uracil DNA glycosylase uses DNA hopping and short-range sliding to trap extrahelical uracils.

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