Amyloid fibrils formed by selective N-, C-terminal sequences of mouse apolipoprotein A-II

Abstract

In mice, amyloidogenic type C apolipoprotein A-II (apoA-II) forms amyloid fibrils in age-associated amyloidosis. To understand the mechanism of amyloid fibril formation by apoA-II, we examined the polymerization of synthetic partial peptides of apoA-II in vitro. None of the partial apoA-II peptides polymerized into amyloid fibrils when tested as a single species mixture. We found a unique mechanism in which N- and C-terminal peptides associated into amyloid fibrils in a 1:1 ratio at pH 2.5. The 11-residue amino acid sequence (6-16), which is a common sequence of type B apoA-II and type C apoA-II proteins in amyloidosis-resistant mice and amyloidosis-susceptible mice, respectively, was critical for polymerization into amyloid fibrils. The 18-residue-long amino acid sequence (48-65) is also necessary for nucleation, but not for the extension phase. These findings suggest that there may be different mechanisms underlying the nucleation and extension phases of apoA-II amyloid fibril formation. We also found that amino acid substitutions between type B apoA-II (Pro5, Val38) and type C apoA-II (Gln5, Ala38) did not affect either phase. The strategy of using synthetic partial peptides of amyloidogenic proteins in vitro is a useful system for understanding amyloid fibril formation and for the development of novel therapies.

Hot spots in apolipoprotein A‐II misfolding and amyloidosis in mice and men

“…We speculate that these factors possibly include the effects of the amino acid substitutions and the pro-peptide on the protein-lipid interactions and on the conformation and dynamics of free apolipoprotein, as proposed in our current structure-based studies of apoA-I [55]. In sum, AmylPred 2 and PASTA consistently predict two hot spots in murine apoA-II, which have been verified experimentally in peptide fragment studies [53,54]. The locations of these hot spots are similar but their predicted amyloid-forming propensities are slightly weaker as compared to their human counterparts (Figs.…”

“…2 and 3), which is higher than in any other exchangeable apolipoprotein explored. This prediction has been verified experimentally for murine apoA-II whose fragments encompassing the predicted segments readily form amyloid in vitro [53,54]. Despite its unusually high sequence propensity, normal apoA-II does not form amyloid in humans.…”

Amyloid-Forming Properties of Human Apolipoproteins: Sequence Analyses and Structural Insights

“…8.4, 8.8, and 8.13). Importantly, the predicted amyloidogenic segments have been verified in the experimental studies of apoC-II, apoA-I, apoA-II and SAA (Wilson et al 2007; Wong et al 2012; Adachi et al 2014; Mendoza-Espinosa et al 2014; Das et al 2014; Sawashita et al 2009; Lu et al 2014). Amyloidogenic segments in many other globular proteins share similar characteristics: they tend to be hydrophobic (with a notable exception of prions), are enriched in aromatic groups, and are often protected from misfolding by forming well-ordered secondary structure such as α-helix (Tzotzos and Doig 2010; Sabate et al 2012).…”

“…8.7). Importantly, the two predicted amyloidogenic segments in apoA-II have been verified experimentally in peptide fragment studies of murine apoA-II that is homologous to the human protein (Sawashita et al 2009). Similar to apoA-I, the amyloidogenic segments in apoA-II are located inside the predicted lipid-binding class-A α-helices (Fig.…”

“…Two major amyloidogenic segments predicted in normal apoA-II have been verified experimentally (Sawashita et al 2009). These segments are located inside the native amphipathic α-helices, and the strongest one (residues 60–70) is predicted to form parallel in-register β-sheet in amyloid.…”