title: Affinity chromatography and capillary electrophoresis for analysis of the yeast ribosomal proteins.
abstract: We present a top down separation platform for yeast ribosomal proteins using affinity chromatography and capillary electrophoresis which is designed to allow deposition of proteins onto a substrate. FLAG tagged ribosomes were affinity purified, and rRNA acid precipitation was performed on the ribosomes followed by capillary electrophoresis to separate the ribosomal proteins. Over 26 peaks were detected with excellent reproducibility (<0.5% RSD migration time). This is the first reported separation of eukaryotic ribosomal proteins using capillary electrophoresis. The two stages in this workflow, affinity chromatography and capillary electrophoresis, share the advantages that they are fast, flexible and have small sample requirements in comparison to more commonly used techniques. This method is a remarkably quick route from cell to separation that has the potential to be coupled to high throughput readout platforms for studies of the ribosomal proteome.
authors.affiliation: The Single Cell Proteomics Group, Institute of Chemical Biology, Department of Chemistry, Imperial College London, South Kensington, London, UK.
authors.forename: Miriam S
authors.initials: MS
authors.lastname: Goyder
authors.forename: Keith R
authors.initials: KR
authors.lastname: Willison
authors.forename: David R
authors.initials: DR
authors.lastname: Klug
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Demello
authors.forename: Oscar
authors.initials: O
authors.lastname: Ces
mesh_terms: D002846:Chromatography, Affinity; D019075:Electrophoresis, Capillary; D005656:Fungal Proteins / Q000032:analysis* / Q000378:metabolism; D020543:Proteome / Q000032:analysis*; D012269:Ribosomal Proteins / Q000032:analysis* / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D005656:Fungal Proteins; D020543:Proteome; D012269:Ribosomal Proteins