Structural basis for poor uracil excision from hairpin DNA

Ghosh, Mahua and Rumpal, Nidhi and Varshney, Umesh and Chary, KVR (2002) Structural basis for poor uracil excision from hairpin DNA. In: European Journal of Biochemistry, 269 (7). pp. 1886-1894.

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Abstract

Two-dimensional NMR and molecular dynamics simulations have been used to determine the three-dimensional structures of two hairpin DNA structures: d-CTAGAG GATCCUTTTGGATCCT (abbreviated as U1-hairpin) and d-CTAGAGGATCCTTUTGGATCCT (abbreviated as U3-hairpin). The (1) H resonances of both of these hairpin structures have been assigned almost completely. NMR restrained molecular dynamics and energy minimization procedures have been used to describe the three-dimensional structures of these hairpins. This study and concurrent NMR structural studies on two other d-CTAGAGGA TCCTUTTGGATCCT (abbreviated as U2-hairpin) and d-CTAGAGGATCCTTTUGGATCCT (abbreviated as U4-hairpin) have shed light upon various interactions reported between Echerichia coli uracil DNA glycosylase (UDG) and uracil-containing DNA. The backbone torsion angles, which partially influence the local conformation of U12 and U14 in U1 and U3-hairpins, respectively, are probably locked in the trans conformation as in the case of U-13 in the U2-hairpin. Such a stretched-out backbone conformation in the vicinity of U-12 and U-14 is thought to be the reason why the K-m value is poor for U1- and U3-hairpins as it is for the U2-hairpin. Furthermore, the bases U-12 and U-14 in both U1- and U3-hairpins adopt an anti conformation, in contrast with the base conformation of U-13 in the U2-hairpin, which adopts a syn conformation. The clear discrepancy observed in the U-base orientation with respect to the sugar moieties could explain why the V-max value is 10- to 20-fold higher for the U1- and U3-hairpins compared with the U2-hairpin. Taken together, these observations support our interpretation that the unfavourable backbone results in a poor K-m value, whereas the unfavourable nucleotide conformation results in a poor V-max value. These two parameters therefore make the U1- and U3-hairpins better substrates for UDG compared with the U2-hairpin, as reported earlier [Kumar, N. V. & Varshney, U. (1997) Nucleic Acids Res. 25, 2336-2343.].

Item Type:	Journal Article
Publication:	European Journal of Biochemistry
Publisher:	John Wiley and Sons
Additional Information:	Copyright of this article belongs to John Wiley and Sons.
Keywords:	hairpin DNA;molecular dynamics;two-dimensional NMR spectroscopy;uracil DNA glycosylase;uracil excision
Department/Centre:	Division of Biological Sciences > Microbiology & Cell Biology
Date Deposited:	28 Jul 2011 06:23
Last Modified:	28 Feb 2019 08:56
URI:	http://eprints.iisc.ac.in/id/eprint/39415

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