Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and mass spectrometry (MS)-based proteomics is useful for characterizing amyloid deposits. We used this approach to analyze transthyretin (TTR) in fat from patients with ATTR amyloidosis associated with various mutations. Extracted proteins were separated by 2D-PAGE. Spots uniquely present in patients were located and investigated by immunoblotting and matrix-assisted laser desorption/ionization time-of-flight MS. Ten patients were studied (4 V30M, 2 E89Q, S50R, G57R, V122I, and E92K). Deposited TTR was always visible as a main train of spots (MW *14 kDa; pI *5.4). In the S50R, G57R and E89Q variants, an additional train shifted in pI was present, containing mutated TTR. MS detected variants as mass shifts in affected peptide ions. All samples but two V30M cases showed C-terminal fragments, with similar migration across mutations. Two-dimensional PAGE-based proteomics provides very characteristic maps in ATTR amyloidosis, confirming the method as a convenient way for molecular dissection of the deposits.
ASJC Scopus subject areas
- Internal Medicine