Solution structure of an antiparallel purine motif triplex containing a T.CG pyrimidine base triple

Triplex formation is an approach of potential use in regulating and mapping of gene sequences. However, such applications have been limited to homogeneous sequences consisting of stretches of purines or pyrimidines. Understanding how heterogeneous duplexes are recognized by a third strand oligonucleotide at the atomic resolution level is an essential step toward broadening the application of triplex formation into biochemical and biomedical areas.

The solution structure of an antiparallel triplex (RRY6) containing a site of inversion (i.e. a T within a homopurine stretch, forming a T.CG base triple) has been determined using NMR-restrained computations in the presence of explicit water. The results reveal that within the RRY6 triplex the conformation of the duplex is mostly B-like and that of the third strand exhibits significant variations in interbase separations and backbone torsion angles. A major displacement of the inversion site T sugar in a 5'-direction, accompanied by the tilt of the T base in T.CG, was observed. The T.CG base triple contains a single hydrogen bond between T O4 and the exposed C amino proton and is stabilized by a number of interstrand and sequential van der Waal contacts. The structural comparisons of RRY6 with two related triplexes indicate localized perturbation at the non-classical base triple site. Various triplexes contain sugars in the C2'-endo family and the global features of their duplexes are similar.

This study provides valuable information concerning the molecular basis of the specific recognition of a Watson-Crick base paired C residue at the inversion sites in the antiparallel triplex and should lead to general rules for designing triplexes containing heterogeneous sequences.