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The vast and intricate connectivity of the human brain combined with the diversity of neurotransmitters are at the root of the combinatorial power that contributes to the functional complexity of the brain. On a molecular level the neurons communicate across the 30 x 10 9 synapses in the human brain by means of neurotransmitters. This synaptic communication is tightly regulated and accordingly alterations in this communication lead to dysfunctions which range from developmental abnormalities to aberrant behavior. Most of the proteins involved in the communication are membrane proteins. The paucity of structural and dynamic information on these proteins has contributed to a lack of functional understanding. In this arena NMR has much to offer. Functional NMR imaging can serve as a bridge between molecular and physiological events in the brain by mapping out tasks, thought patterns, memory, etc. in exquisite anatomical detail. Moreover, diffusion sensitive NMR imaging can characterize electrical current flow in biological tissue. Recent advances in both solution and solid-state NMR demonstrate the potential of these spectroscopic approaches for solving membrane protein structure, one of the major challenges in structural biology and neuroscience today. A great many processes important to biological function are mediated by molecules integrated in, or attached to, membrane surfaces. Many of the molecules involved in these processes are membrane proteins, but they also include lipids and various glycoconjugates that adorn membrane surfaces with distinct collections of complex carbohydrates. Structural and dynamic characterizations of these macromolecules have been achieved infrequently due to the heterogeneous nature of the samples. Bilayers form an inherently heterogeneous environment with a hydrophobic interior and a hydrophilic exterior, in addition the oligosaccharides decorating the membrane surfaces are themselves heterogeneous. Such complexity makes crystallization very difficult and NMR spectra complicated. But recently, the first glycoprotein structures have been solved and much progress has been made in developing the technology necessary to characterize membrane protein structure and dynamics. The development of orientational constraints is central to this latter technology. These structural constraints define how individual sites are oriented with respect to a global molecular axis. In addition, the development of liquid crystalline media has been very important. Small aggregates of lipid mixtures (bicelles) orient cooperatively in a magnetic field to produce an alignment media for NMR spectroscopy. These preparations are now being used in both solution NMR to gain residual dipolar couplings for water soluble proteins as structural constraints and in solid-state NMR where they take full advantage of the nuclear spin interaction anisotropy. The discoveries of the utility of orientational data and magnetic field oriented media are examples of the types of discoveries that can be anticipated through research at high magnetic fields. The expectation that well in excess of 1000 new potential drugs
in the field of neuroscience will be derived from genomics and
that more than 90% of these will be targeted to channels and receptors,
exemplifies the importance of structure, dynamics, function correlations
for characterizing the drug binding pockets. Among the several
classes of important membrane associated proteins, ligand gated
channels, such as the acetylcholine and NMDA receptors are being
actively studied. Low resolution structural information on the
acetylcholine receptor is available from electron microscopy.
Recent solid-state NMR of lamellar phase lipid preparation and
solution NMR of micellar preparations of the 25 residue M2 Studies of To meet the challenges of characterizing such protein structure and dynamics not only will high magnetic fields be necessary, but advances in NMR technology to gain sensitivity and resolution are desperately needed. Developments in solution NMR at very high field strengths leading to improved resolution for much larger molecular weight species will bring the full potential of solution NMR to a much greater array of proteins than previously possible. Optically pumped He and Xe have great potential for enhancing MR images. Dynamic nuclear polarization as described earlier has demonstrated approximately 200 fold sensitivity enhancement in frozen aqueous preparations. With such large signal enhancements it should be possible to dramatically improve resolution in solid-state spectra via higher dimensional experiments, reduced sample size requirements, and/or dramatically shorten signal acquisition times. Recent advances in probe and pulse sequence technology for solid-state NMR at 800 MHz have led to substantially decreased linewidths in magic angle spinning spectra. |
peptide is leading to higher resolution structural insights.
This peptide is one of the membrane bound helices from the acetylcholine
receptor that displays electrophysiological properties similar
to that of the protein. By using a combination of solution and
solid-state NMR spectroscopy to study the same peptide in membrane
mimetic environments substantial progress has been made in showing
that their structure corresponds to 
-helices that are aligned in the lipid bilayer at an angle of
12° with respect to the bilayer normal. Most importantly, these
studies set the stage for investigations of larger segments, including
the entire membrane associated portion of these proteins, when
high field magnets become available. The goal is to be able to
determine the full three-dimensional structures of large membrane
proteins by NMR spectroscopy, and the success of this endeavor
will be strongly influenced by access to very high field magnets.
-amyloid, the putative cause of Alzheimer's disease represents
another example of a heterogeneous structure where solid-state
NMR has made a unique contribution. By using magic angle spinning
methods it is possible to work with unoriented samples. Through
distance measurements with homonuclear rotational resonance experiments,
the first structure of an aggregating protein, a 9-mer derived
from the C-terminal end of