Spectroscopic Probes of Protein Structure
With few exceptions, the goal of most protein purification efforts is to obtain a sample that is not only pure, but that also maintains the protein in its native (i.e., biologically active) conformation. The ability to describe the conformation of a protein in solution, and to relate changes in conformation with biological activity, is thus a major focus of protein science. The most detailed description of protein structures come from the determination of the complete three-dimensional arrangement of protein components in space, from x-ray crystallographic or nuclear magnetic resonance (NMR) studies. Despite their power, however, these methods are not without their attendant drawbacks. X-ray diffraction studies of proteins are dependent on obtaining protein crystals of sufficient size and quality to yield usable diffraction patterns. This can often be a time consuming, and not necessarily successful, undertaking. Even when high quality crystals are obtained, solving the structure from the resulting diffraction patterns is a laborious and time consuming effort. Add to this the fact that certain classes of proteins, such as integral membrane proteins, are inherently difficult to crystallize, and one soon realizes that x-ray crystallography, while an extremely powerful method, is not a panacea for protein structural problems. Multidimensional NMR spectroscopy likewise suffers from certain difficulties that restrict its utility. Perhaps the greatest limit to the use of NMR spectroscopy for solving protein structures is that the complexity of the multidimensional data is such that the size of a protein that can reasonably be solved is limited to about 100 amino acids or so. While significant efforts are currently being put forth to push up the size limit for NMR spectroscopy, at least for the present this method is limited to relatively small proteins.
KeywordsCircular Dichroic Circular Dichroic Spectrum Difference Spectrum Tryptophan Residue Nuclear Magnetic Resonance Spectroscopy
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