Paramagnetic additives, with their effect on spectral linewidths, have also been used in EPR spectroscopy. Additionally, the orientation of a peptide within a membrane can be obtained by the anisotropic hyperfine tensor of a rigidly attached nitroxide label. Besides these magnetic resonance techniques a series of other methods to probe the orientation of peptides in membranes has been developed, consisting of fluorescence-, infrared- and oriented circular dichroism spectroscopy, colorimetry, interface-sensitive X-ray and neutron scattering and Quartz crystal microbalance. Membrane-bound proteins and peptides constitute a major class of all expressed proteins and peptides of a genome and are involved in crucial biological processes. Statistical analysis of several genomes of different organisms indicated that 30% of the encoded proteins and peptides are membrane-bound. Many naturally occurring peptides bind to biological membranes, examples include peptide hormones, bacterial toxins and antimicrobial peptides with the latter being probably the largest and best characterized group of them. A variety of either linear or cyclic antimicrobial peptides have been described to date and they often provide a first unspecific defense mechanism against microbial invasion. Many of these peptides are believed to target microbial cytosolic cell membranes, disrupting them upon interaction. Besides knowing the structure, deciphering the mode of interaction with membranes or membrane-mimetics is necessary to understand their function.
While the target of many antimicrobial peptides is the microbial lipid bilayer itself, membrane-binding peptide hormones and bacterial toxins likely act on proteins located in the membrane. Besides membrane-bound peptides, which can be structurally characterized easily by solution NMR spectroscopy, there are an enormous number of membrane-binding proteins. Although the 3D structure determination of soluble proteins by X-ray crystallography and NMR spectroscopy is well-established, far fewer structures of membrane-bound proteins have been solved to date.
Structural studies of membrane proteins are considerably complicated by their amphipathic or hydrophobic nature and resulting solubility problems in aqueous buffers. Less than 2% of all structures in the Protein Data Bank (PDB) are membrane-bound proteins although they account for about 30% of all expressed proteins.įrequently, their limited stability in the absence of lipids constitutes an additional problem. Noteworthy, about 45% of all pharmaceuticals target G-protein coupled receptors, which are also a group of membrane-bound proteins. The limited structural information available for these proteins stems either from difficulties in crystallization or their large size, resulting in broad signals and limited spectral dispersion in NMR spectra.
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