posted on 2016-02-20, 22:18authored byWilliam
M. Tay, Jennifer G. Bryant, Patricia K. Martin, A. Jeremy Nix, Bernadette M. Cusack, Terrone L. Rosenberry
Endogenous amyloid-β (Aβ) oligomeric aggregates
have
been proposed as toxic agents in Alzheimer’s disease (AD).
Knowledge of their structures not only may provide insight into the
basis of their neurotoxicities but also may reveal new targets for
therapeutic drugs and diagnostic tools. However, the low levels of
these Aβ oligomers have impeded structural characterization.
Evidence suggests that the endogenous oligomers are covalently modified
in vivo. In this report, we demonstrate an established mass spectrometry
(MS) methodology called precursor ion mapping (PIM)
that potentially may be applied to endogenous oligomer characterization.
First, we illustrate the use of this PIM technique with a synthetic
Aβ(1–40) monomer sample that had been cross-linked with
transglutaminase (TGase) and digested with pepsin. From PIM analysis
of an Aβ(4–13) MS/MS fragment, precursor ions were identified
that corresponded to peptic fragments of three TGase cross-linked
species: Aβ(4–19)--(4–19), Aβ(4–19)--(20–34),
and Aβ(1–19)--(20–34). Next, we demonstrate the
applicability of the PIM technique to an endogenous Aβ sample
that had been purified and concentrated by immunoaffinity chromatography.
Without pepsin digestion, we successfully identified the full length
and C-terminally truncated monomeric Aβ species 1–35
to 1–42, along with select methionine-oxidized counterparts.
Because PIM focuses only on a subpopulation of ions, namely the related
precursor ions, the resulting spectra are of increased specificity
and sensitivity. Therefore, this methodology shows great promise for
structural analysis and identification of post-translational modification(s)
in endogenous Aβ oligomers.