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Amyloid fibrils derived from the apolipoprotein A1 Leu174Ser variant contain elements of ordered helical structure

¹ Department of Biochemistry, University of Pavia, 27100 Pavia, Italy
² Centro per la Studio e la Cura delle Amiloidosi, 27100 Pavia, Italy
³ Department of Biochemistry, University of Cambridge, CB2 1GA, UK
⁴ Laboratory of Biotechnology, IRCCS Policlinico S. Matteo, 27100 Pavia, Italy
⁵ Dipartimento di Scienze e Tecnologie Biomediche, Università di Udine, 33100 Udine, Italy
⁶ Centro Grandi Strumenti, Università di Pavia, 27100 Pavia, Italy

Reprint requests to: Vittorio Bellotti, Dipartimento di Biochimica, via Taramelli 3/b Università di Pavia, 27100 Pavia, Italy; e-mail: vbellot{at}unipv.it; fax: 390382-423108.

We recently described a new apolipoprotein A1 variant presenting a Leu174Ser replacement mutation that is associated with a familial form of systemic amyloidosis displaying predominant heart involvement. We have now identified a second unrelated patient with very similar clinical presentation and carrying the identical apolipoprotein A1 mutation. In this new patient the main protein constituent of the amyloid fibrils is the polypeptide derived from the first 93 residues of the protein, the identical fragment to that found in the patient previously described to carry this mutation. The X-ray fiber diffraction pattern obtained from preparations of partially aligned fibrils displays the cross-ß reflections characteristic of all amyloid fibrils. In addition to these cross-ß reflections, other reflections suggest the presence of well-defined coiled-coil helical structure arranged with a defined orientation within the fibrils. In both cases the fibrils contain a trace amount of full-length apolipoprotein A1 with an apparent prevalence of the wild-type species over the variant protein. We have found a ratio of full-length wild-type to mutant protein in plasma HDL of three to one. The polypeptide 1–93 purified from natural fibrils can be solubilized in aqueous solutions containing denaturants, and after removal of denaturants it acquires a monomeric state that, based on CD and NMR studies, has a predominantly random coil structure. The addition of phospholipids to the monomeric form induces the formation of some helical structure, thought most likely to occur at the C-terminal end of the polypeptide.

Keywords: Amyloidosis; apolipoprotein A1; proteolysis; fibrillogenesis

Abbreviations: TIC, total ion current • ESI-MS, electrospray ionization mass spectrometry • apoA1, apolipoprotein A1 • POPC, palmitoyl-oleoyl phosphatidylcholine • CD, circular dichroism • HDL, high density lipoprotein • NOE, nuclear Overhauser effect • TOCSY, total correlation spectroscopy • NOESY, NOE spectroscopy • 1D, one dimensional • 2D, two dimensional • SAP, serum amyloid-P component.

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