title: Desalting electroeluted proteins with hydrophilic interaction chromatography.
abstract: We describe a chromatographic procedure for the removal of sodium dodecyl sulfate (SDS) from proteins isolated by electroelution. It involves chromatography of electroeluates on poly(2-hydroxyethyl-aspartamide) silica, a support initially developed for hydrophilic interaction chromatography. The electroeluate, dialyzed against ammonium bicarbonate-SDS buffer, is directly injected onto the column, which is equilibrated in an n-propanol concentration greater than 60%. Bound proteins are eluted with a gradient of decreasing n-propanol. This procedure removes essentially all of the Coomassie blue-related contaminants and separates SDS from the protein. Due to the use of volatile buffer systems, the proteins are recovered in completely salt-free form, which facilitates further protein manipulation. After removal of the organic solvent from the chromatographic desalting step, the recovered proteins are directly amenable to N-terminal protein sequencing and, after evaporation of the organic phase, to enzymatic digestions and subsequent separation of fragments by reverse-phase HPLC.
authors.affiliation: Department of Biochemistry, University of Basel, Switzerland.
authors.forename: P
authors.initials: P
authors.lastname: Jenö
authors.forename: P E
authors.initials: PE
authors.lastname: Scherer
authors.forename: U
authors.initials: U
authors.lastname: Manning-Krieg
authors.forename: M
authors.initials: M
authors.lastname: Horst
mesh_terms: D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D011506:Proteins / Q000302:isolation & purification*; D012967:Sodium Dodecyl Sulfate / Q000302:isolation & purification*
chemical_list: D011506:Proteins; D012967:Sodium Dodecyl Sulfate