Ion Mobility-Mass Spectrometry Fuels Chiral Peptide Chemistry
Chiral inversion of amino acids is thought to modulate the structure and function of amyloid beta (Aβ) but these processes are poorly understood. Herein the authors develop an ion mobility-mass spectrometry approach to study chirality-regulated structural features of Aβ fragments.
In this work, we present an innovative concept and potentially new drug target for Alzheimer’s disease (AD) therapy by investigating chiral effects on amyloid-beta (Aβ) via a novel multidimensional ion mobility-mass spectrometry platform. As chiral Aβ peptides with partial amino acid D-isomerization have been detected in AD brains, there is a possibility that D-isomerized Aβ play a vital role in early AD pathogenesis and development. However, since Aβ D-isomerization is age-dependent and is present at low stoichiometry (e.g. less than 10%), the role of chiral Aβ has long been ignored and largely underexplored, in part due to lack of effective tools.
Figure 1. Newly-proposed concept about AD development and a novel analytical platform based on in-solution kinetics analysis and gas-phase multi-dimensional ion mobility-mass spectrometry that allows dissecting effect of chiral chemistry involved in AD. D-isomerized Aβ could be a new target due to its distinct monomer structure, oligomerization behavior and receptor recognition feature. The method, iCAP, is abbreviated from integrated chirality anatomy platform.
In this study, we develop and establish an innovative analytical platform based on ion mobility mass spectrometry (IM-MS) (Figure 1), we discover and characterize the important role of chirality in regulating AD-related Aβ self-assembly and receptor recognition. Benefiting from these rational designs that target Aβ chiral chemistry, distinct structural and molecular differences have been revealed between wild type and D-isomerized Aβ, including its monomer structure, oligomerization behavior and its receptor-recognition and binding characteristics. In addition to the crosstalking effects among those epimeric Aβ during oligomerization, the differential contributions of the chirality of Aβ N-terminal and C-terminal fragments were also interrogated, suggesting cooperative effects. The current results could facilitate future investigations of novel therapeutic treatments for AD as new insights can be obtained via elucidation of the roles of D-isomerized Aβ in early AD development, diagnosis, and prognosis.
The innovation of the work presented in this manuscript can be summarized below:
- 1)First report on a multifaceted approach to simultaneously provide molecular and structural evidence for chiral effect of Aβ.
- 2)Development of a broadly applicable and rationally designed chiral anatomy platform.
- 3)Discovery of differential structural roles of chiral Aβ N-terminal and C-terminal fragments and novel molecular findings that suggest the possibility of D-isomerized Aβ as a potentially new AD drug target.
The key idea of chiral amplification through metal binding was originally inspired by previous reports including our own research experience on zinc finger peptide-zinc binding (J. Am. Soc. Mass Spectrom. (2017) 28: 2658-2664) and other IM-MS-based peptide-metal binding studies (J. Am. Soc. Mass Spectrom. (2017) 28: 1293-1303). These previous studies suggested that metal binding can enhance structural difference. We therefore choose copper as our candidate metal, as it has its unique isotopic distribution that helps us identify the binding events much easier even only with MS1 measurements. Besides, Cu2+ binds to most peptides with a moderate to high affinity and thus we can capture such types of binding in the gas phase even after desolvation. Lastly, it is also widely involved in many real biological situations which may link our experiments biologically to some real disease or human health case. The next question, however, comes to how to maximize the chiral amplification power and how to quantitatively characterize/report such chiral amplification. Gongyu recalled reading a paper from Analytical Chemistry (Anal. Chem. 2016, 88, 2335−2344), entitled “Multidimensional Analysis of 16 Glucose Isomers by Ion Mobility Spectrometry” where the authors demonstrated a multidimensional data visualization method for analyte isomer analysis with data collected from an IM-MS platform. Gongyu was excited to adopt this conception, but with significant modifications, including more rational choice of individual coordinates/vectors, namely, the collisional cross-sections (CCS) for zero-, one- and two-copper-bound peptide species. These three CCSs are highly dependent on the metal-binding events and thus the chiral effects can be evaluated. The next step was to plot each of these 3D vectors into a 3D scattering space. Furthermore, after discussing with Dr. Xin Peng from Lingjun Li Research Group, Gongyu decided to use the spatial distance in this 3D scattering space to quantitatively characterize the D/L Structural Difference (DLSD). The DLSD method has now been proven very useful in this kind of evaluations. We also adopted the same key concept for Aβ oligomer chiral amplification and quantitative characterization.
From the initial submission to final acceptance of our revised manuscript for publication in Nature Communications, the reviewers and editor have been helpful to continuously improve this work. Currently in the Li lab, we are extending the fragment-based in vitro study to full-length Aβ in vivo exploration, starting with some in cell tests. Notably, we anticipate combining this iCAP conception with our previous D-amino acid residue localization method published in Analytical Chemistry (Anal. Chem. 2014, 86, 2972−2981), entitled “Site-Specific Characterization of D‑Amino Acid Containing Peptide Epimers by Ion Mobility Spectrometry”. This combined study can contribute to a systematic interrogation of chiral inversion of various amino acid residues on full-length Aβ stereoisomers and while allowing precise localization of the D-amino acid residues. Ultimately, we hope to provide more concrete molecular basis for potentially new AD drug targets, including the previously ignored structural differences caused by amino acid chirality.