Deoxyribonucleic acids can form a wide variety of structural motifs which differ greatly from the typical antiparallel duplex stabilized by Watson-Crick base pairing. Many of these structures are thought to occur in vivo and may have essential roles in the biology of the cell. Among these is the parallel-stranded duplex-a structural motif in which DNA strands associate in a head-to-head fashion with the 5' ends at the same end of the duplex-which is stabilized by reverse Watson-Crick base pairing. In this study, parallel- and antiparallel-stranded DNA duplexes formed from two different 12-mer oligonucleotides were studied using native electrospray ionization combined with trapped ion mobility spectrometry and mass spectrometry. The DNA duplex charge plays an important role in the gas-phase mobility profile, with a more compact form in negative mode than in positive mode (Delta Omega approximate to 100 angstrom(2) between -4 and +4). Despite sequence mismatches, homo- and hetero-DNA duplexes were formed in solution and transfer to the gas phase, where a more compact structure was observed for the parallel compared to the antiparallel duplexes (Delta Omega approximate to 50 angstrom(2)), in good agreement with theoretical calculations. Theoretical studies suggest that a reduction (or compaction) along the helical axis of the parallel and antiparallel DNA duplexes is observed upon transfer to the gas phase.