Sequential NMR assignments of labile protons in DNA using two-dimensional nuclear-overhauser-enhancemnt spectroscopy with three jump-and-return pulse sequences

Abstract

Two-dimensional nuclear Overhauser enhancement (NOESY) spectra of labile protons were recorded in H₂O solutions of a protein and of a DNA duplex, using a modification of the standard NOESY experiment with all three 90° pulses replaced by jump-and-return sequences. For the protein as well as the DNA fragment the strategically important spectral regions could be recorded with good sensitivity and free of artifacts. Using this procedure, sequence-specific assignments were obtained for the imino protons, C2H of adenine, and C4NH₂ of cytosine in a 23-base-pair DNA duplex which includes the 17-base-pair O_R3 repressor binding site of bacteriophage λ. Based on comparison with previously published results on the isolated O_R3 binding site, these data were used for a study of chain termination effects on the chemical shifts of imino proton resonances of DNA duplexes.