  While studying nucleic acids to reveal the weak interactions responsible for
their three-dimensional structure and for their interactions with drugs, we
also contributed to the field of biomolecular mass spectrometry, both in terms
of fundamental understanding and with new methodological developments. A first
goal was to develop mass spectrometry approaches to detect non-covalent
interactions between antitumor drugs and their DNA target. Twenty years ago,
our attention turned towards specific DNA structures such as the G-quadruplex
(a structure formed by guanine-rich strands). Mass spectrometry allows to
discern which molecules interact with one another by measuring the masses of
the complexes, and quantity the affinities by measuring their abundance. The
most important findings came from unexpected masses. For example, we showed the
formation of higher- or lower-order structures by G-quadruplexes used in
traditional biophysical assays. We can also derive complete thermodynamic and
kinetic description of G-quadruplex folding pathways by measuring cation
binding, one at a time. Getting quantitative information required accounting
for nonspecific adduct formation and for the response factors of the different
molecular forms. But with these caveats in mind, the approach is mature enough
for routine biophysical characterization of nucleic acids. A second goal is to
obtain more detailed structural information on each of the complexes separated
by the mass spectrometer. One such approaches is ion mobility spectrometry, and
even today the challenge lies in the structural interpretation of the
measurements. We showed that, although structures such as G-quadruplexes are
well-preserved in the MS conditions, double helices actually get more compact
in the gas phase. These major rearrangements forced us to challenge comfortable
assumptions. Further work is still needed to generalize how to deduce
structures in solution from ion mobility spectrometry data, and in particular
how to account for the electrospray charging mechanisms and for ion internal
energy effects. These studies also called for complementary approaches to ion
mobility. Recently, we applied isotope exchange labeling mass spectrometry to
characterize nucleic acid structures for the first time, and we reported the
first ever circular dichroism ion spectroscopy measurement on mass-selected
trapped ions. Circular dichroism plays a key role in assigning the stacking
topology, and our new method now opens the door to characterizing a wide
variety of chiral molecule by mass spectrometry. In summary, advanced mass
spectrometry approaches to characterize gas-phase structures work well for
G-quadruplexes because they are stiffened by inner cations. The next objective
will be to generalize these methodologies to a wider range of nucleic acid
structures.
