Biomedical Engineering Reference
In-Depth Information
16.7
Conclusion
The past decade has seen an explosion in the number of membrane-protein struc-
tures determined by liquid-state NMR spectroscopy, especially those with multiple
transmembrane-spanning
-helices. This progress came as a result of recent innova-
tions in sample preparation, strategies for isotopic labeling, developments in NMR
pulse sequences, and new alignment media. Alignment is an essential component of
contemporary protein NMR studies, especially for membrane proteins whose highly
asymmetric structures are dominated by
'
-sheets and are difficult to
characterize based on short-range distance or intramolecular angle measurements.
The emergence of the first detergent-resistant liquid crystals based on DNA-origami
nanostructures has facilitated the accurate measurement of RDCs on membrane
protein, from which global orientation and structural information can be derived.
In addition, the new method introduced recently by Chou et al
.
on the UCP2
carrier, which combines orientation restraints derived from samples weakly aligned
in a DNA-nanotube liquid crystal (RDCs) and semiquantitative distance restraints
from paramagnetic-relaxation-enhancement measurements (PRE), opens a route to
extend the size limit of solution-NMR-based de novo structure determination of
membrane proteins. Through the structural analysis of different functional states
of membrane proteins, this new method can potentially open new horizons in
biochemical research because of the large number of helical membrane proteins
of great biomedical interest between the sizes of 20 and 30 kDa. Over the next
10 years, we are likely to witness many more NMR structures from the huge
number of membrane proteins yet to be characterized. The structures obtained so
far have highlighted the benefits of combining structural DNA nanotechnology
and liquid-state NMR spectroscopy for understanding the structure and function of
membrane proteins. Now that advancement in atomic resolution structural studies
for membrane protein are rapidly gaining momentum, the innovative focus of the
field is shifting to improve and develop experimental systems that can provide
structural information on the arrangement of functional domains relative to one
another, and to gain insights into how this may be altered during signaling. While
the DNA-nanotube technology is expected to be extensively used for a wide
variety of membrane proteins, it will be important to facilitate the measurement
of linearly independent restraints to achieve more structural information. To this
end, additional DNA-nanostructure-based alignment media will be needed. Due to
the tunability of DNA nanostructure's shape, size, and chemical functionalization,
opportunities exist for improvements to be made in intrinsic ordering of the liquid
crystals and in compatibility of the liquid crystals with positively charged proteins.
DNA nanostructures with different shapes and charge distributions may induce
linearly independent alignment tensors in target proteins, which would facilitate the
collection of additional independent sets of structural restraints. Beyond solving
the three-dimensional structures using RDC patterns and NOE approaches, a
large variety of developments in well-established methods based on RDC data
'
-helices or
“
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