H-1-NMR STUDIES OF THE 5-(HYDROXYMETHYL)-2'-DEOXYURIDINE CONTAINING TF1 BINDING-SITE

The pyrimidine base 5-(hydroxymethyl)-2'-deoxyuridine (HmU) is a commo n nucleotide in SPO1 phage DNA. Numerous transcriptional proteins bind HmU-containing DNA preferentially implicating a regulatory function o f HmU. We Rave investigated the conformation and dynamics of d-(5'-CHm UCHmUACACGHmUGHmUA-GAG-OH-3')(2) (HmU-DNA). This oligonucleotide mimic s the consensus sequence of Transcription Factor 1 (TF1). The HmU-DNA was compared to the thymine-containing oligonucleotide. NOESY and DQF COSY spectroscopy provided resonance assignments of nonexchangeable an d exchangeable protons, intranucleotide, internucleotide acid intrastr and proton-proton distances, and dihedral angle constraints. Methylene protons of the hydroxymethyl group are nonequivalent protons and the hydroxymethyl group is not freely rotating. The hydroxymethyl group ad opts a specific orientation with the OH group oriented on the 3' side of the plane of the base, Analysis of imino proton resonances and NOEs indicates additional end base pair fraying and a temperature-induced transition to a conformation in which the internal HmU-A base pairs ar e disrupted or have reduced lifetimes. Orientation of the hydroxymethy l group indicates the presence of internucleotide intrastrand hydrogen bonding between the HmU12C5 hydroxyl group and A13, All sugars in bot h DNAs show a C2'endo conformation (typical of B-DNA).