Insights from ion mobility-mass spectrometry, infrared spectroscopy, and molecular dynamics simulations on nicotinamide adenine dinucleotide structural dynamics: NAD⁺vs.NADH

Abstract

Nicotinamide adenine dinucleotide (NAD) is found in all living cells where the oxidized (NAD) and reduced (NADH) forms play important roles in many enzymatic reactions. However, little is known about NAD and NADH conformational changes and kinetics as a function of the cell environment. In the present work, an analytical workflow is utilized to study NAD and NADH dynamics as a function of the organic content in solution using fluorescence lifetime spectroscopy and in the gas-phase using trapped ion mobility spectrometry coupled to mass spectrometry (TIMS-MS) and infrared multiple photon dissociation (IRMPD) spectroscopy. NAD solution time decay studies showed a two-component distribution, assigned to changes from a "close" to "open" conformation with the increase of the organic content. NAD gas-phase studies using nESI-TIMS-MS displayed two ion mobility bands for NAD protonated and sodiated species, while four and two ion mobility bands were observed for NADH protonated and sodiated species, respectively. Changes in the mobility profiles were observed for NADH as a function of the starting solution conditions and the time after desolvation, while NAD profiles showed no dependence. IRMPD spectroscopy of NAD and NADH protonated species in the 800-1800 and 3200-3700 cm spectral regions showed common and signature bands between the NAD forms. Candidate structures were proposed for NAD and NADH kinetically trapped intermediates of the protonated and sodiated species, based on their collision cross sections and IR profiles. Results showed that NAD and NADH species exist in open, stack, and closed conformations and that the driving force for conformational dynamics is hydrogen bonding of the N-H-O and O-H-O forms with ribose rings.

Identification of Lasso Peptide Topologies Using Native Nanoelectrospray Ionization-Trapped Ion Mobility Spectrometry–Mass Spectrometry

“…In particular, the recent introduction of trapped IMS (TIMS) and its integration with MS analyzers, − has found multiple applications in analytical workflows with high mobility resolving power ( R up to ∼400) in a compact geometry . TIMS-MS devices have proven useful for rapid separation and structural elucidation of biomolecules, ,− for screening, and targeted , analysis of complex mixtures, tracking isomerization kinetics, − characterizing the conformational spaces of peptides, separation of d -amino acid containing peptides, DNA, proteins, − and macromolecular complexes in native and denatured states …”

Single‐stranded DNA structural diversity: TAGGGT from monomers to dimers to tetramer formation

“…The TIMS cell was operated using a fill/trap/ramp/wait sequence of 14.5/.15/100–500/.765 ms. A constant 880 kHz 200 Vpp rf was applied to all electrodes including the entrance funnel, the TIMS analyzer section and the exit funnel (for schematic, see Figure S1, supporting information). Ions were introduced using low‐energy conditions (i.e., V Def = ±60 V, V cap = ±50 V, and V fun = 0 V) to avoid ion activation prior to the mobility analysis . CCS (Ω) values were calculated from reduced mobility (K 0 ) values using the Mason‐Schamp equation: where z is the charge of the ion, K B is the Boltzmann constant, T is the temperature, N* is the bath gas number density and m I and m b are the masses of the molecular ion and the bath gas, respectively.…”

Vibrational signatures of curcumin’s chelation in copper(II) complexes: An appraisal by IRMPD spectroscopy

“…[22][23][24] Based on a photofragmentation process triggered by the sequential absorption of resonant IR photons, 25 in recent years, IRMPD spectroscopy has become a pivotal tool to shed light on molecular features and binding interactions of a variety of (de)protonated (in)organic ions, [26][27][28][29] metal-and halide-bound adducts of (modified) amino acids and peptides, [30][31][32][33][34][35][36][37] nucleobases and nucleotides, [38][39][40][41][42][43] saccharides, [44][45][46] metabolites, 47,48 and cofactors. 49,50 This tool is employed here to expand our information on curcumin's scavenging of neurotoxic Cu(II) metal ions in the prospect to relate direct structural features to biological activity.…”