Biomedical Engineering Reference
In-Depth Information
channels using predominantly information from solid-state NMR studies of
oriented membranes. 4,5 Also, in this work, mechanistically important subtle
kinks and ionisation states of residues in trans-membrane peptides have been
described.
Here we outline some of the major solid-state NMR methods and their
applications that have become established over recent years, as well as several
newer developments likely to drive methods forward in the future.
13.2 Samples
Many early MAS studies of proteins focussed on small microcrystalline
samples 6-10 which usually give rise to very narrow linewidths because of their
high degree of conformational homogeneity. Coupled with the fact that the
proteins chosen were usually relatively robust with respect to sample
conditions (especially temperature), these samples have formed very useful
model systems.
Amyloid fibrils, which are difficult to study at atomic resolution by any other
technique, were another early target for structural studies of proteins. 11-15
However, the spectral quality varies quite considerably from protein to protein 16
or between different fibril morphologies. 17 Conformational heterogeneity or a
high degree of dynamics often leads to broad resonance lines, 14,15 and for some
proteins the spectral quality has only been sufficient for the determination of
models rather than full structures. 15 A notable exception are the HET-s fibrils
which give rise to very high-quality spectra and whose structure was determined
to high resolution. 18 The HET-s fibrils appear to form a relatively stable
structure which may be due to the fact that fibrils are the native state for HET-s
fibrils, i.e., the protein may have evolved to form more rigid and stable fibrils
compared
to
many
other
protein
fibrils
which
are
generally
a
result
of
misfolding.
Membrane proteins form a large target for solid-state NMR studies since
they can be conducted in the native lipid environment 3,19 in contrast to
solution NMR and X-ray crystallography investigations. Static solid-state
NMR experiments using oriented bilayer samples have been carried out for
many years and have yielded many structures, especially for small, membrane-
embedded single-helix peptides. 5 Structural and functional models are now
also being described using solid-state MAS NMR data. For smaller proteins,
such as phospholamban 20 and the influenza virus A M2 ion channel 4 full
structure determinations have been possible. Many assignments have been
reported for larger membrane proteins, but no ab initio structures have been
determined to date. Several of these studies have focussed on the relatively
robust (bacterial) rhodopsin family of proteins. 21-25 Other proteins studied
include the membrane-embedded enzyme DsbB, 26 outer membrane porin G
(OmpG), 27 the ArtMP ATP-binding-cassette (ABC) transporter 28 and a
voltage dependent anion channel 1 (VDAC1). 29 Combining both naturally
crystalline (2D) arrays of a protein with labelling strategies, NMR crystal-
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