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Publisher Summary The chapter discusses the structural analysis of amyloid fibrils by electron microscopy and X-ray diffraction. A method to isolate amyloid fibrils from tissue was developed, and such preparations of purified fibrils exhibited the classic staining and birefringent characteristics of crude amyloid samples. The further use of high-resolution electron microscopy, examining ultrathin sections of tissue and isolated material, confirmed the fibrillar form as the characteristic appearance of amyloid in the electron microscope. With the ability to prepare samples of isolated amyloid fibrils it began to be possible to probe the molecular structure of the fibrils using X-ray diffraction. The initial use of this technique produced diffraction patterns that showed that the fibrils were composed of polypeptide chains, extended in the so-called cross-β-conformation. Subsequent analyses by X-ray diffraction on a variety of amyloids and also by NMR analysis have confirmed that the protein chains in all amyloid fibrils are predominantly in the cross-β-conformation. The structures of the soluble, globular forms of several amyloidogenic proteins have been determined by the single crystal X-ray crystallography. The origins of the different amyloidoses appear quite diverse, with some arising from a genetic mutation and others apparently from polypeptide misprocessing or from an unusual accumulation of full-length wild-type protein. |
