NMR-derived solution structure of a 17mer hydroxymethyluracil-containing DNA

Abstract

Incorporation of 5-(hydroxymethyl)-2'-deoxyuridine into DNA in place of thymine by SPO1, a Bacillus subtilis bacteriophage, allows the viral DNA to bind selectively to transcription factor 1. We have synthesized a TF1-binding site: d(5'-ACCHACHCHHHGHAGGT-3')-d(5'-ACCHACAAAGAGHAGGT-3') and studied this molecule using NMR spectroscopy. The chemical shifts of exchangeable and non-exchangeable protons were sequentially assigned. Absence of corresponding NOEs in the imino-imino region suggested that the end base pairs did not form Watson-Crick hydrogen bond. Restrained molecular dynamics calculation yielded a family of B-DNA structures whose r.m.s.d. was 0.66 Å (all atoms) for the internal 15 bp. The helical twist was 38.5°per step. The base pairs were situated directly on the helix axis (X-displacement = -0.2 Å). All sugars exhibited C2'-endo puckering with P = 167.3°and υ(max) = 38.2°. The OH groups of all hmU bases resided on the 3' side of the base plane and may affect the base orientation relative to the sugar plane as the average χ value for all hmU was 4°more positive than that of other nucleosides (258°versus 254°). Positive roll angles (p) and small flanking twists to) at hmU suggested that the two hmU-A base pair steps open toward the minor grooves.