PXD004867

PXD004867 is an original dataset announced via ProteomeXchange.

Title	The human amyloid beta peptide interactome
Description	The amyloid beta (Aβ) peptide represents a 37 to 49 amino acids endoproteolytic fragment of the amyloid precursor protein. The cellular biology that governs the formation and clearance of Aβ has been understood to play a critical role in Alzheimer’s disease (AD). The primary objective of this study was to generate an in-depth inventory of human brain proteins that oligomeric preparations of Aβ1-42 can bind to using an unbiased in vitro discovery approach. Synthetic Aβ1-42 peptides and a brain extract generated from adult human frontal lobe tissue served in these studies as baits and biological source material, respectively. oAβ1-42 was prepared by aggregating the peptide at 4 ºC for 24 h, using previously described procedures known to generate amyloid-β-derived diffusible ligands (ADDLs). Because the interaction with a given binding partner may rely on a binding epitope that comprises N- or C-terminal residues of Aβ1-42, two separate experiments (I and II) were conducted, which differed in the orientation the oAβ1-42 bait was tethered to the affinity matrix. To facilitate meaningful comparisons across experiments, the method of Aβ1-42 capture was not based on immunoaffinity reagents. Instead, alternative Aβ1-42 baits were equipped with biotin moieties attached to the N- or C-terminus by a 6-carbon linker chain, enabling their consistent affinity-capture on streptavidin agarose matrices. Biotin-saturated streptavidin agarose matrices served as negative controls and three biological replicates of samples and controls were generated for each of the three separate interactome datasets by reproducing the affinity-capture step side-by-side on three separate streptavidin agarose affinity matrices that had been saturated with the biotinylated baits. To identify differences in protein-protein interactions of monomeric versus oligomeric Aβ1-42, a 3rd interactome experiment was conducted in which oAβ1-42-biotin or mAβ1-42-biotin served as baits. Digitonin-solubilized brain extracts, which are known to comprise extracellular and most cellular proteins (except for nuclear proteins) served as biological starting material, consistent with the main subcellular areas previously reported to harbor Aβ. Following extensive washes of affinity matrices in their protein-bound state, binders to the bait peptides were eluted by rapid acidification, fully denatured in 9 M urea, and trypsinized. To avoid notorious confounders related to variances in the subsequent handling and analysis of samples, individual peptide mixtures were labeled with distinct isobaric tandem mass tags (TMT) in a six-plex format, then combined and concomitantly subjected to ZipTip-based pre-analysis clean-up by strong cation exchange (SCX) and reversed phase (RP) separation. Four-hour split-free reversed phase nanospray separations were online coupled to an Orbitrap Fusion Tribrid mass spectrometer, which was configured to run an MS3 analysis method.
HostingRepository	PRIDE
AnnounceDate	2017-07-06
AnnouncementXML	Submission_2017-07-19_00:05:33.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Declan Williams
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	S-pyridylethyl-L-cysteine; phosphorylated residue; monohydroxylated residue; deamidated residue
Instrument	Orbitrap Fusion ETD

Revision	Datetime	Status	ChangeLog Entry
0	2016-08-19 01:38:41	ID requested
1	2017-07-05 09:10:52	announced
2	2017-07-06 00:30:21	announced	Updated project metadata.
⏵ 3	2017-07-19 00:05:35	announced	Updated project metadata.

Wang H, Muiznieks LD, Ghosh P, Williams D, Solarski M, Fang A, Ruiz-Riquelme A, Pom, è, s R, Watts JC, Chakrabartty A, Wille H, Sharpe S, Schmitt-Ulms G, peptide and favors the formation of distinct oligomers. Elife, 6():(2017) [pubmed]

curator keyword: Biological, Biomedical

submitter keyword: Alzheimer’s disease, amyloid beta, human, brain, frontal lobe, affinity purification, orbitrap

Gerold Schmitt-Ulms
contact affiliation	University of Toronto
contact email	g.schmittulms@utoronto.ca
lab head
Declan Williams
contact affiliation	University of Toronto
contact email	gwillym.williams@utoronto.ca
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD004867

PRIDE project URI

• PRIDE
  1. PXD004867
     1. Label: PRIDE project
     2. Name: The human amyloid beta peptide interactome