Key Segments of N-Glycans in the Gas Phase
A R T I C L E S
entropy gain associated with its higher flexibility favors its
population at the initial elevated temperature of the vaporization
source and the structure is frozen in the early stages of the free
jet expansion. Relaxation into the global potential energy
minimum would be inhibited if the barrier to conversion from
B16 to A16 was much higher than the energy of the collisions
experienced in the expansion. The opposite situation was met
in benzyl lactoside16 where the minimum potential energy
structure, a rigid conformer with two strong inter-ring H bonds,
was stabilized in the free jet expansion, although free energy
calculations suggested that it was not favored at high temper-
atures.
In contrast to the R(1,3) disaccharide, comparisons between
the IR spectra of the two principal conformers A16 and B16 and
those of its constituent monosaccharide units do not suggest
clear correspondences. Each of the suggested conformational
assignments includes a mannose subunit in which the OH groups
are arranged in a counterclockwise arrangement, as in RæMan
B, and a comparison of the calculated and experimental IR
spectra associated with A16 and B16 does indicate a clear
connection between the vibrational bands observed at 3694 cm-1
(indicated by a wedge in Figure 5c,d) and the corresponding
band in the IR spectrum of RæMan B: both are associated with
the σ3′ stretching mode. A correspondence with the other bands
of the spectrum of conformer B of RæMan is also suggested
(indicated by the dotted lines in Figure 5), but the congested
IRID spectra and the uncertainties in their detailed assignments,
particularly of the conformer B16, make comparisons difficult.
ing band absorbs at a high wavenumber, 3690 cm-1. In larger
oligosaccharides, more severe IR spectral congestion can be
anticipated, but it will be concentrated mainly in the intermediate
spectral region lying between 3600 and 3650 cm-1, where intra-
ring H bonded OH groups absorb. The high and low wave-
number ends of the spectrum should remain much clearer and
continue to provide reliable indicators of the conformational
families to which the conformers belong.
The spectral features of the isolated monosaccharide residues
may also be used to refine the assignment of larger oligosac-
charide structures. This provides a more general supplementary
building block approach to the study of complex structures,
based upon an alphabet of established IR spectral signatures of
different conformations of the monosaccharide unitswhen their
spectroscopic patterns are retained. If they are not retained, their
evolution can be understood by analyzing the modification of
the hydrogen bonded networks in the disaccharides, for example,
the retention (or disruption) of the secondary structural motifs
generated by intraresidue H-bonding. The concept of secondary
structure for carbohydrates has not been addressed very clearly
yet. The spectral analysis of their IR spectra can be eased by
considering each monosaccharide unit in a larger carbohydrate
structure, as a module with or without interactions promoted
by inter-ring H bonds.
Finally, feedback from the increasing body of experimental
data will help to inform and guide future theoretical confor-
mational searches. It is possible that using IR spectra of subunits
of a large system to understand its own vibrational spectroscopy
and guide the fine-tuning of the computational conformational
identification will meet some limitations due to spectral conges-
tion as the size of the system increases, but the computational
time bottleneck will probably occur much earlier. If the
conformational search can be informed by experiment to include
only those geometries that incorporate features clearly identified
by the experimental spectrum, it should be possible to extend
further the limits of the current strategy. This will prove essential
for defining the intrinsic conformational preferences of larger
oligosaccharides going toward the core pentasaccharide of
N-linked glycans.
Conclusion
Despite the challenge in tackling such a complicated con-
formational landscape, there is good reason to be confident about
the coarse-grained structural assignments, based upon the
qualitative analysis of the observed spectral signatures, the match
between the observed and calculated spectra associated with
the low-lying conformers, their relative energies, and where
possible, comparisons between the spectra of the disaccharide
conformers and their monosaccharide components. Conforma-
tional assignments as definite and precise as those that can be
obtained for smaller molecules are beyond the scope of current
strategies, but there is no doubt about the distinction between
the two conformational families now identified: those with, A13
and A16, or without, B16, a (single) inter-ring hydrogen bond.
The ability to separate and assign such structures experimentally
will help in identifying the regions of larger oligosaccharides
that can provide some rigidity, or a source of flexibility, or
targets for the binding of structural water molecules.11
The general IR spectral signatures that have been character-
ized, as well as identifying membership of broad conformer
families, can also identify noninteracting, free OH groups that
are potentially important sites for interaction with the environ-
ment.9,11 The spectrum of conformer A13 in ManR(1,3)Manæ
provides an excellent example. There is an inter-ring hydrogen
bond, linking OH2 to O6′, which shifts the IR absorption of its
stretching mode to 3520 cm-1, but OH6′ does not interact with
any other group in the moleculeswhich makes it an ideal
binding site for solvent molecules11sand the associated stretch-
Acknowledgment. We are grateful for the encouragement
provided by Drs. Mark Wormald and David Chambers and the
financial and material support provided by the EPSRC, the
Royal Society (R.A.J. USA Research Fellowship; L.C.S.
University Research Fellowship), the Leverhulme Trust (Grant
F/08788D), the CLRC Laser Loan Pool, and the Physical and
Theoretical Chemistry Laboratory at Oxford.
Supporting Information Available: Detailed description of
the synthesis and characterization of 1 and 2. Cartesian
coordinates and total energies of the conformations assigned to
A13, A16, and B16. Complete refs 16 and 18. Larger scale display
of the conformations of 1 and 2 shown in Figures 4 and 7. This
material is available free of charge via the Internet at
JA055891V
9
J. AM. CHEM. SOC. VOL. 128, NO. 6, 2006 1981