Analysis of x-ray diffraction patterns from amyloid of biopsied vitreous humor and kidney of transthyretin (TTR) Met30 familial amyloidotic polyneuropathy (FAP) patients: axially arrayed TTR monomers constitute the protofilament

Abstract

Familial amyloidotic polyneuropathy (FAP) is characterized by deposits of amyloid jbers in which the major protein component is transthyretin (TTR). Nearly fifty mutations have been reported for the TTR in hereditary FAP. Protein crystallography of mutant TTRs has shown that the molecular structures of the variant molecules are similar to those found in the wild type. On this basis, the FAP fibers were initially proposed to consist of native-like TTR tetramers. In the current paper, we used x-ray fiber diffraction to study the structure of FAP fibers from biopsy samples of vitreous humor and kidney. The reflections of the vitreous sample showed a cross-β diffraction pattern. All the meridional reflections were indexed by a one-dimensional, 29 Å-period lattice, and the equatorial reflections were indexed by an apparent one-dimensional 67 Å-period lattice. The x-ray intensity distribution indicated that the unit structure, which is similar to a TTR monomer, is composed of a pair of β-sheets consisting of four hydrogen-bonded β-chains per sheet, with the β-chains oriented approximately normal to the fiber axis. The axial disposition of these units, with a 29 Á-period, constitutes the protofilament; and a tetrameric lateral assembly of the protofilaments containing the core domain of the ∼20 Å-wide β-sheet structure constitutes the FAP amyloid fiber. An inter-fiber separation of 75 Å in these concentrated samples accounts for the apparent one-dimensional lattice perpendicular to the fiber axis. In the delipidated kidney FAP sample, the diffraction pattern indicated a pair of β-sheets, suggesting that the protofilament structure in kidney is similar to that in vitreous humor. In the non-delipidated sample the successive sharp reflections indexed to a one-dimensional, 48.9 Å-lattice, and the electron density projection showed a density elevation at the center of a lipid bilayer. This suggests that lipid may be associated with the monomeric TTR in the kidney FAP protofilament.

Key Words:

• amyloid
• fiber diffraction
• β-pleated sheet
• fibrillogenesis
• familial amyloidotic polyneuropathy
• amyloidosis