Amyloid fibril formation and seeding by wild-type human lysozyme and its disease-related mutational variants

Wild-type human lysozyme and its two stable amyloidogenic variants have bee n found to form partially folded states at low pH. These states are charact erized by extensive disruption of tertiary interactions and partial loss of secondary structure. Incubation of the proteins at pH 2.0 and 37 degrees C (Ile56Thr and Asp67His variants) or 57 degrees C (wild-type) results in th e formation of large numbers of fibrils over several days of incubation. Sm aller numbers of fibrils could be observed under other conditions, includin g neutral pH. These fibrils were analyzed by electron microscopy, Congo red birefringence, thioflavine-T binding, and X-ray fiber diffraction, which u nequivocally show their amyloid character. These data demonstrate that amyl oidogenicity is an intrinsic property of human lysozyme and does not requir e the presence of specific mutations in its primary structure. The amyloid fibril formation is greatly facilitated, however, by the introduction of "s eeds" of preformed fibrils to the solutions of the variant proteins, sugges ting that seeding effects could be important in the development of systemic amyloidosis. Fibril formation by wild-type human lysozyme is greatly accel erated by fibrils of the variant proteins and vice versa, showing that seed ing is not specific to a given protein. The fact that wild-type lysozyme ha s not been found in ex vivo deposits from patients suffering from this dise ase is likely to be related to the much lower population of incompletely fo lded states for the wild-type protein compared to its amyloidogenic variant s under physiological conditions. These results support the concept that th e ability to form amyloid is a generic property of proteins, but one that i s mitigated against in a normally functioning organism. (C) 2000 Academic P ress.