Familial Alzheimer's disease mutations differentially alter amyloid β-protein oligomerization

Abstract

Although most cases of Alzheimer's disease (AD) are sporadic, ∼5% of cases are genetic in origin. These cases, known as familial Alzheimer's disease (FAD), are caused by mutations that alter the rate of production or the primary structure of the amyloid β-protein (Aβ). Changes in the primary structure of Aβ alter the peptide's assembly and toxic activity. Recently, a primary working hypothesis for AD has evolved where causation has been attributed to early, soluble peptide oligomer states. Here we posit that both experimental and pathological differences between FAD-related mutants and wild-type Aβ could be reflected in the early oligomer distributions of these peptides. We use ion mobility-based mass spectrometry to probe the structure and early aggregation states of three mutant forms of Aβ40 and Aβ42: Tottori (D7N), Flemish (A21G), and Arctic (E22G). Our results indicate that the FAD-related amino acid substitutions have no noticeable effect on Aβ monomer cross section, indicating there are no major structural changes in the monomers. However, we observe significant changes to the aggregation states populated by the various Aβ mutants, indicating that structural changes present in the monomers are reflected in the oligomers. Moreover, the early oligomer distributions differ for each mutant, suggesting a possible structural basis for the varied pathogenesis of different forms of FAD.

Figure 1

Mass spectrum of Aβ40 D7N, the Tottori mutant. Peaks corresponding to z/n = −4, −3, −2, and −5/2 are present, where z is the charge and n is the oligomer size. Mass spectra of all other alloforms considered here are given in Supporting Information, Figure S1.

Figure 2

Plots of the z/n = −3 ATD of (a) wt Aβ40 and (b) Aβ40 D7N. Two overlapping peaks represent a gas phase structure (○) and dehydrated solution structure (□). See ref (28) for a detailed discussion of this assignment. A peak at ∼500 μs in the Aβ40 D7N ATD represents a larger assembly. The red peaks are theoretical representations of a single monomer isomer at the indicated arrival time (see eq 3).

Figure 3

Arrival time distributions of z/n = −5/2 for (a) wt Aβ40 and (b) wt Aβ42 at an injection voltage of 35 V, where z is the charge and n is the oligomer size. ATDs of z/n = −5/2 are also given for (c,e) Aβ40 D7N and (d,f) Aβ42 D7N at injection voltages of 35 and 100 V, respectively. The narrow red peaks are calculated from eq 3 and are the width of a single isomer.

Figure 4

Arrival time distributions of z/n = −5/2 for the Flemish mutant (A21G) of (a) Aβ40 and (b) Aβ42 and the Arctic mutant (E22G) of (c) Aβ40 and (d) Aβ42, where z is the charge and n is the oligomer size. All ATDs were recorded with an injection voltage of 35 V. The narrow red peaks are calculated from eq 3 and are the width of a single isomer.

Scheme 1. Mechanisms of Early Oligomer Formation of (a) Aβ40 wt, (b) Aβ40 A21G, (c) Aβ40 D7N, and (d) Aβ40 E22G

Normally, wt Aβ40 forms monomer, dimer, and tetramer, where the structure of the tetramer is closed. The D7N and E22G mutations allow the peptide to form larger oligomers like a hexamer and even, in the case of E22G, a dodecamer. Of interest is the fact that the A21G mutant forms an open tetramer but no hexamer suggesting a different kind of interfacial bonding for this alloform.

Scheme 2. Mechanisms of Early Oligomer Formation of (a) Aβ42 wt, (b) Aβ42 D7N or Aβ42 E22G, and (c) Aβ42 A21G

D7N and E22G mutations are shown together due to the similarity of their mechanisms. Normally, wt Aβ42 forms monomer through dodecamer. The D7N and E22G mutations increase the amount of dodecamer formed. The A21G mutation stops Aβ42 oligomerization at the hexamer. Of more importance, the hexamer structue for this alloform is open, not a planar cylic ring like all other hexamers we observe in the other alloforms. Hence ring stacking is not available to it, and no dodecamer is formed along with decrease protofibril and fibril formation.