title: Structures of the kinetically trapped i-motif DNA intermediates.
abstract: In the present work, the conformational dynamics and folding pathways of i-motif DNA were studied in solution and in the gas-phase as a function of the solution pH conditions using circular dichroism (CD), photoacoustic calorimetry analysis (PAC), trapped ion mobility spectrometry-mass spectrometry (TIMS-MS), and molecular dynamics (MD). Solution studies showed at thermodynamic equilibrium the existence of a two-state folding mechanism, whereas during the pH = 7.0 → 4.5 transition a fast and slow phase (ΔHfast+ ΔHslow= 43 ± 7 kcal mol-1) with a volume change associated with the formation of hemiprotonated cytosine base pairs and concomitant collapse of the i-motif oligonucleotide into a compact conformation were observed. TIMS-MS experiments showed that gas-phase, kinetically trapped i-motif DNA intermediates produced by nanoESI are preserved, with relative abundances depending on the solution pH conditions. In particular, a folded i-motif DNA structure was observed in nanoESI-TIMS-MS for low charge states in both positive and negative ion mode (e.g., z = ±3 to ±5) at low pH conditions. As solution pH increases, the cytosine neutralization leads to the loss of cytosine-cytosine+(C·CH+) base pairing in the CCC strands and in those conditions we observe partially unfolded i-motif DNA conformations in nanoESI-TIMS-MS for higher charge states (e.g., z = -6 to -9). Collisional induced activation prior to TIMS-MS showed the existence of multiple local free energy minima, associated with the i-motif DNA unfolding at z = -6 charge state. For the first time, candidate gas-phase structures are proposed based on mobility measurements of the i-motif DNA unfolding pathway. Moreover, the inspection of partially unfolded i-motif DNA structures (z = -7 and z = -8 charge states) showed that the presence of inner cations may or may not induce conformational changes in the gas-phase. For example, incorporation of ammonium adducts does not lead to major conformational changes while sodium adducts may lead to the formation of sodium mediated bonds between two negatively charged sides inducing the stabilization towards more compact structures in new local, free energy minima in the gas-phase.
authors.affiliation: Department of Chemistry and Biochemistry, Florida International University, Miami, USA. fernandf@fiu.edu.
authors.forename: Alyssa
authors.initials: A
authors.lastname: Garabedian
authors.affiliation: Department of Chemistry and Biochemistry, Florida International University, Miami, USA. fernandf@fiu.edu.
authors.forename: David
authors.initials: D
authors.lastname: Butcher
authors.affiliation: The RNA Institute, University at Albany, Albany, USA.
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Lippens
authors.affiliation: Department of Chemistry and Biochemistry, Florida International University, Miami, USA. fernandf@fiu.edu and Biomolecular Science Institute, Florida International University, Miami, USA.
authors.forename: Jaroslava
authors.initials: J
authors.lastname: Miksovska
authors.affiliation: Biomolecular Science Institute, Florida International University, Miami, USA and Department of Physics, Florida International University, Miami, USA.
authors.forename: Prem P
authors.initials: PP
authors.lastname: Chapagain
authors.affiliation: The RNA Institute, University at Albany, Albany, USA.
authors.forename: Daniele
authors.initials: D
authors.lastname: Fabris
authors.affiliation: Bruker Daltonics Inc., Billerica, Massachusetts, USA.
authors.forename: Mark E
authors.initials: ME
authors.lastname: Ridgeway
authors.affiliation: Bruker Daltonics Inc., Billerica, Massachusetts, USA.
authors.forename: Melvin A
authors.initials: MA
authors.lastname: Park
authors.affiliation: Department of Chemistry and Biochemistry, Florida International University, Miami, USA. fernandf@fiu.edu and Biomolecular Science Institute, Florida International University, Miami, USA.
authors.forename: Francisco
authors.initials: F
authors.lastname: Fernandez-Lima
mesh_terms: D002151:Calorimetry; D002942:Circular Dichroism; D003596:Cytosine / Q000737:chemistry; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D000075663:Ion Mobility Spectrometry; D007700:Kinetics; D056004:Molecular Dynamics Simulation; D009690:Nucleic Acid Conformation; D059372:Nucleotide Motifs; D021241:Spectrometry, Mass, Electrospray Ionization; D013816:Thermodynamics
chemical_list: D003596:Cytosine; D004247:DNA