Elucidating the mechanism of familial amyloidosis-Finnish type: NMR studies of human gelsolin domain 2

Familial amyloidosis-Finnish type (FAF) results from a single mutation at r esidue 187 (D187N or D187Y) within domain 2 of the actin-regulating protein gelsolin. The mutation somehow allows a masked cleavage site to be exposed , leading to the first step in the formation of an amyloidogenic fragment. We have performed NMR experiments investigating structural and dynamic chan ges between wild-type (WT) and D187N gelsolin domain 2 (D2). On mutation, n o significant structural or dynamic changes occur at or near the cleavage s ite. Areas in conformational exchange are observed between beta-strand 4 an d alpha-helix 1 and within the loop region following beta-strand 5. Chemica l shift differences are noted along the face of cu-helix 1 that packs onto the p-sheet, suggesting an altered conformation. Conformational changes wit hin these areas can have an effect on actin binding and may explain why D18 7N gelsolin is inactive. {H-1-N-15} nuclear Overhauser effect and chemical shift data suggest that the C-terminal tail of D187N gelsolin D2 is less st ructured than WT by up to six residues. In the crystal structure of equine gelsolin. the C-terminal tail of D2 lies across a large cleft between domai ns 1 and 2 where the masked cleavage site sits. We propose that the D187N m utation destabilizes the C-terminal tail of D2 resulting in a more exposed cleavage site leading to the first proteolysis step in the formation of the amyloidogenic fragment.