Solution structure of a truncated anti-MUC1 DNA aptamer determined by mesoscale modeling and NMR

Mucin 1 is a well-established target for the early diagnosis of epithelial cancers. The nucleotides of the S1.3/S2.2 DNA aptamer involved in binding to variable number tandem repeat mucin 1 peptides have been identified using footprinting experiments. The majority of these binding nucleotides are located in the 25-nucleotide variable region of the total aptamer. Imino proton and 2D NMR spectra of truncated and total aptamers in supercooled water reveal common hydrogen-bonding networks and point to a similar secondary structure for this 25-mer sequence alone or embedded within the total aptamer. NMR titration experiments confirm that the TTT triloop structure is the primary binding site and show that the initial structure of the truncated aptamers is conserved upon interaction with variable number tandem repeat peptides. The thermal dependence of the NMR chemical shift data shows that the base-paired nucleotides melt cooperatively at 47 +/- 4 degrees C. The structure of the 25-mer oligonucleotide was determined using a new combined mesoscale molecular modeling, molecular dynamics and NMR spectroscopy investigation. It contains three WatsonCrick pairs, three consecutive mispairs and four WatsonCrick pairs capped by a TTT triloop motif. The 3D model structures (PDB ) and biopolymer chain elasticity molecular models are consistent with both NMR and long unconstrained molecular dynamics (10 ns) in explicit water, respectively.