Characterization of the Amyloid Fibril as a Cross- Protein

Abstract

The major component of amyloid is a fibrous protein (1) with a unique organization as seen by high resolution electron microscopy (2). A second component, a pentagonal rod, has been isolated from amyloid deposits and demonstrated to be identical with a circulating a globulia (P-component) (3). This component is distinct from the fibril in its ultrastructure, chemistry, and immunological interactions and constitutes only a minute part of amyloid deposits. In 1960 we obtained an X-ray diffraction powder pattern of an amyloid-rich tissue. In subsequent years improved methods of isolation of amyloid were devised and X-ray diffraction patterns of these more purified preparations were obtained. Results were presented that indicated the presence of a cross-β pattern. The present study was carried out on amyloid fibrils isolated by various techniques to determine whether this molecular structure was consistently present in all preparations, including those freed from the pentagonal unit. Meterials and Methods Amyloid fibrils were obtained from the spleens of patients with primary, secondary, and myelomaassociated amyloidosis and prepared by the following methods: (1) homogenization of whole amyloid-laden spleen, centrifugation and recovery of a top-layer rich in amyloid fibrils (4); (2) sucrose gradient separation of the top-layer specimen to obtain a purer preparation (5); (3) reprecipitation of amyloid fibrils after they had been solubilized in dilute alkali at pH 11.5 (6); (4) a “water-soluble” amyloid prepared after purifying amyloid-laden spleen by exhaustive saline washes followed by homogenization in water (7)