Dataset Information

Cardiac Atrial Compartmentalisation Proteomics: A Modified Density Gradient Method to Analyse Endo-lysosomal Proteins

ABSTRACT: Evidence that lysosomes are important in cardiac physiology and pathology is now emerging. We describe a label-free method suitable for small cardiac tissue biopsies based on a density-separated fractionation protocol, which allows us to study endo-lysosomal (EL) proteins in guinea pig atria. The three most relevant density gradient fractions obtained were analysed using Western Blot, enzyme activity assay and MS/MS peptide analysis (adapted discontinuous Percoll, and sucrose differential density gradient). These fractions comprise whole tissue lysate; sarcoplasmic reticulum (SR), mitochondrial proteins (Mito) (1.3 g/mL); and ‘pure’ EL with negligible contamination from SR or Mito (1.04 g/mL). Kyoto Encyclopedia of Genes and Genomes, Reactome, Panther and Gene Ontology pathway analysis showed good coverage of RAB proteins, lysosomal cathepsins (including cardiac-specific cathepsin D) in the purified EL fraction. The most significant EL proteins recovered included catalytic activity proteins. We thus present a comprehensive protocol and dataset of guinea-pig atrial EL focussed organelle proteomics

INSTRUMENT(S): Q Exactive

ORGANISM(S): Cavia Porcellus (guinea Pig)

TISSUE(S): Heart

SUBMITTER: Georgina Berridge

LAB HEAD: Rebecca A B Burton

PROVIDER: PXD021277 | Pride | 2021-09-09

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	FL984_MSQ1439_20180902_RebeccaCapel_1.mgf	Mgf
	FL984_MSQ1439_20180902_RebeccaCapel_1.mzxml	Mzxml
	FL984_MSQ1439_20180902_RebeccaCapel_1.pep.xml	Pepxml
	FL984_MSQ1439_20180902_RebeccaCapel_1.raw	Raw
	FL984_MSQ1439_20180902_RebeccaCapel_2.mgf	Mgf

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A modified density gradient proteomic-based method to analyze endolysosomal proteins in cardiac tissue.

Ayagama Thamali T Bose Samuel J SJ Capel Rebecca A RA Priestman David A DA Berridge Georgina G Fischer Roman R Galione Antony A Platt Frances M FM Kramer Holger H Burton Rebecca A B RAB

iScience 20210804 9

The importance of lysosomes in cardiac physiology and pathology is well established, and evidence for roles in calcium signaling is emerging. We describe a label-free proteomics method suitable for small cardiac tissue biopsies based on density-separated fractionation, which allows study of endolysosomal (EL) proteins. Density gradient fractions corresponding to tissue lysate; sarcoplasmic reticulum (SR), mitochondria (Mito) (1.3 g/mL); and EL with negligible contamination from SR or Mito (1.04 ...[more]

PMID: 34466782

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