Dataset Information

Novel Peptidomic Approach for Identification of Low and High Molecular Weight Tauopathy Peptides Following Calpain Digestion, and Primary Culture Neurotoxic Challenges.

ABSTRACT: Tauopathy is a class of a neurodegenerative disorder linked with tau hyperphosphorylation, proteolysis, and aggregation. Tau can be subjected to proteolysis upon calpain activation in Alzheimer disease (AD), and traumatic brain injury (TBI). We and others have extensively researched calpain-mediated tau breakdown products (Tau-BDP; 45K, 35K, and 17K). Tau proteolysis might also generate low molecular weight (LMW ?10K) proteolytic peptides after neurodegenerative damage. In this study, we have subjected purified tau protein (phospho and non-phospho) and mouse brain lysate to calpain-1 digestion to characterize the LMW generated by nano-liquid chromatography coupled to electrospray ionization to tandem mass spectrometry (nano-LC-ESI-MS/MS). We have also challenged differentiated primary cerebrocortical neuronal cultures (CTX) with neurotoxic agents (calcium ionophore calcimycin (A23187), staurosporine (STS), N-methyl-D-aspartate (NMDA), and Maitotoxin (MTX)) that mimic neurodegeneration to investigate the peptidome released into the conditioned cell media. We used a simple workflow in which we fractionate LMW calpain-mediated tau peptides by ultrafiltration (molecular weight cut-off value (MWCO) of 10K) and subject filtrate fractions to nano-LC-MS/MS analysis. The high molecular weight (HMW) peptides and intact proteins retained on the filter were analyzed separately by western blotting using total and phospho-specific tau antibodies. We have identified several novel proteolytic tau peptides (phosphorylated and non-phosphorylated) that are only present in samples treated with calpain or cell-based calpain activation model (particularly N- and C-terminal peptides). Our findings can help in developing future research strategies emphasizing on the suppression of tau proteolysis as a target.

SUBMITTER: Yadikar H

PROVIDER: S-EPMC6829287 | biostudies-literature | 2019 Oct

REPOSITORIES: biostudies-literature

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Novel Peptidomic Approach for Identification of Low and High Molecular Weight Tauopathy Peptides Following Calpain Digestion, and Primary Culture Neurotoxic Challenges.

Yadikar Hamad H Johnson Connor C Pafundi Niko N Mouhawasse Edwin E Nguyen Lynn L Torres Isabel I Kurup Milin M Yang Zhihui Z Kobeissy Firas F Yost Richard R Wang Kevin K KK

International journal of molecular sciences 20191021 20

Tauopathy is a class of a neurodegenerative disorder linked with tau hyperphosphorylation, proteolysis, and aggregation. Tau can be subjected to proteolysis upon calpain activation in Alzheimer disease (AD), and traumatic brain injury (TBI). We and others have extensively researched calpain-mediated tau breakdown products (Tau-BDP; 45K, 35K, and 17K). Tau proteolysis might also generate low molecular weight (LMW ≤10K) proteolytic peptides after neurodegenerative damage. In this study, we have su ...[more]

PMID: 31640160

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Project description:Protein misfolding disorders are associated with conformational changes in specific proteins, leading to the formation of potentially neurotoxic amyloid fibrils. During pathogenesis of prion disease, the prion protein misfolds into ?-sheet rich, protease-resistant isoforms. A key, hydrophobic domain within the prion protein, comprising residues 109-122, recapitulates many properties of the full protein, such as helix-to-sheet structural transition, formation of fibrils and cytotoxicity of the misfolded isoform. Using all-atom, molecular simulations, it is demonstrated that the monomeric 109-122 peptide has a preference for ?-helical conformations, but that this peptide can also form ?-hairpin structures resulting from turns around specific glycine residues of the peptide. Altering a single amino acid within the 109-122 peptide (A117V, associated with familial prion disease) increases the prevalence of ?-hairpin formation and these observations are replicated in a longer peptide, comprising residues 106-126. Multi-molecule simulations of aggregation yield different assemblies of peptide molecules composed of conformationally-distinct monomer units. Small molecular assemblies, consistent with oligomers, comprise peptide monomers in a ?-hairpin-like conformation and in many simulations appear to exist only transiently. Conversely, larger assemblies are comprised of extended peptides in predominately antiparallel ?-sheets and are stable relative to the length of the simulations. These larger assemblies are consistent with amyloid fibrils, show cross-? structure and can form through elongation of monomer units within pre-existing oligomers. In some simulations, assemblies containing both ?-hairpin and linear peptides are evident. Thus, in this work oligomers are on pathway to fibril formation and a preference for ?-hairpin structure should enhance oligomer formation whilst inhibiting maturation into fibrils. These simulations provide an important new atomic-level model for the formation of oligomers and fibrils of the prion protein and suggest that stabilization of ?-hairpin structure may enhance cellular toxicity by altering the balance between oligomeric and fibrillar protein assemblies.

Dataset Information

Novel Peptidomic Approach for Identification of Low and High Molecular Weight Tauopathy Peptides Following Calpain Digestion, and Primary Culture Neurotoxic Challenges.

Publications

Novel Peptidomic Approach for Identification of Low and High Molecular Weight Tauopathy Peptides Following Calpain Digestion, and Primary Culture Neurotoxic Challenges.

Similar Datasets

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