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A last type of effect of misfolded protein is seen in amyloid diseases; in which protein deposits in the cells as fibrils. A number of common diseases of old age, such as Alzheimer's disease fit into this category, and in some cases an inherited version occurs, which has enabled study of the defective protein.
One such example is transthyretin, a protein that transports thyroxin and retinol binding protein in the bloodstream and cerebrospinal fluid. In senile systemic amyloidosis, which affects  people over 80, transthyretin forms fibrillar deposits in the heart. which leads to congestive heart failure. An inherited version, Familial amyloid polyneuropathy (FAP) affects much younger people; causing protein deposits in the heart, and in many other tissues; deposits around nerves can lead to paralysis. FAP patients are mostly heteozygous for the transthyretin gene, and over 50 single site mutations are known.
    The structure of wild type TTR was determined in the 1970's by Colin.Blake and his colleagues; the biological unit is a tetramer. Each monomer has two 4- stranded b-sheets, and a short a -helix. Anti-parallel b -sheet interactions link monomers into dimers and a short loop from each monomer forms the main dimer-dimer interaction. These pairs of loops keep the two halves of the structure apart forming an internal channel.
Here are two views of the tetramer

Side view

Top view

The internal channel has two binding sites for thyroxin. Only a small proportion of the plasma TTR molecules are involved in thyroxin transport, a much higher proportion are found tightly bound to retinol-binding protein (RBP); the function of this is thought to be to stabilize the binding of retinol to RBP, keeping it in the plasma by reducing its filtration from the blood by the kidneys. Retinol is made from Vitamin A, and is important in regulation of vertebrate development.

The structure of several FAP TTR variants is known, and they are virtually identical to the normal form .To examine the stucture of transthyretin and the mutants interactively the following links will allow you to see the stuctures using Rasmol or Chime. The pdb files from expasy can be seen with Rasmol if you  have it installed, and from the new PDB (Protein Data Bank) site at RCSB there are several options available (click on view structure, and choose from VRML, Chime, or Rasmol). The following links will allow you to see the original wild-type stucture 2pab, from [expasy] or [RCSB], the later (refined) 1tta from [expasy] or [RCSB]; and the mutant forms 1ttb (Ala109 to Thr) [expasy][RCSB] and 1ttc (Val30 to Met) [expasy][RCSB]. The PDB files of other mutants (such as 1tsh,1ttr), and complexes of transthyretin with retinol binding protein, retinoic acid, thyroxine and various inhibitors are also available from expasy,RCSB., or the PDB mirror site at EBI. 
What then causes this protein whose normal form is a small ovoid molecule to make long insoluble fibrils?
There is not yet conclusive evidence, but from the evidence below it seems likely that the monomer must be able to adopt an alternative conformation that forms long b-strands. Jeffery Kelly has found that if wild type TTR is denatured at low pH (5.1-3.9) the tetramers are dissociated, and amyloid forms.((The FAP TTR variants make much more amyloid than the wild type and at higher pH. He proposed that this occurs at physiological pH in the mutant forms of the protein, and involves the degradation pathway, because the pH of lysosomes is in the range where mutant protein forms fibrils. Blake and colleagues have found that the crystal structure of the Met30 variant when crystallised in isomorphous form to native TTR, had a shift in the position of Cys 10, the only cysteine in the subunit, making it slightly more exposed. This suggested that it is possible that the linear aggregates are held together by disulphide bonds to cysteine residues on adjacent subunits. ( A study by N.Schormann and co-workers found no significant differences between the structures of  3 mutants that formed amyloid, and 2 that did not by X-ray crystallography, but they found a large amount of truncated protein, missing the first 48 amino acids, in the mutants prone to amyloid. They suggest that the mutants have a amino acid that causes the area around position 48 to be more exposed and therefore more prone to proteolysis and that the truncated protein can then not form a stable tetramer. ( Study of the fibrils is difficult because of its insolubility making NMR solution studies impossible and they do not make good crystals. However Blake and Serpell () have analyzed fibrils from one FAP patient homozygous for the mutation substituting Met for the normal Val at position 30 by X-ray diffraction, using partially dried samples analyzed with a synchrotron radiation source, and found a pattern consistent with a long b -helical structure, with 24 b -strands per turn of the b -helix.


Model of a helical b-sheet structure

Four twisted b -helices make up a proto-filament (50-60A), and four of these associate to form a fibril as seen in electron microscopy (130A).

() Model of a fibril made up of 4 fibrils

Recently T. Klabunde and colleagues have studied the binding of thyroxine, and of some anti-inflammatory drugs, that inhibit fibril formation in vitro, to transthyretin. They showed that the binding site for thyroxine has 3 pairs of small depressions. Two of these on each monomer bind to iodines of thyroxine, in two alternative conformations, and the bound thyroxine stabilises the tetramer. They have designed some drugs that bind more strongly  than thyroxine, which should therefore stabilise the tetrameric form, so there is now hope of effective treatment for SSA and FAP. ().

Pedro Soares 2002 | Updated: 10/15/2002