title: High-yield two-step chromatographic procedure for purification of fatty acid-binding protein from human heart.
abstract: A high-yield procedure for the purification of cytoplasmic fatty acid-binding protein from human heart (H-FABP) is described. H-FABP was purified by gel permeation chromatography on a Sephacryl S-200 column followed by anion-exchange chromatography on a Sepharose Q fast-flow column at pH 7.0. At this pH H-FABP binds strongly to the column and can be selectively eluted with a salt gradient. The two-step procedure showed a high degree of reproducibility. On average 50 mg of H-FABP was obtained from 150 g of human heart tissue, which corresponds to a recovery of about 50%. Purity was confirmed by gel electrophoresis and isoelectric focusing. Binding of oleic acid to purified H-FABP, using the Lipidex 1000 assay, revealed a maximal binding of 0.75 +/- 0.01 mol fatty acid/mol protein and a dissociation constant of 0.19 +/- 0.01 microM.
authors.affiliation: Department of Physiology, University of Limburg, Maastricht, Netherlands.
authors.forename: F A
authors.initials: FA
authors.lastname: van Nieuwenhoven
authors.forename: M M
authors.initials: MM
authors.lastname: Vork
authors.forename: D A
authors.initials: DA
authors.lastname: Surtel
authors.forename: A H
authors.initials: AH
authors.lastname: Kleine
authors.forename: G J
authors.initials: GJ
authors.lastname: van der Vusse
authors.forename: J F
authors.initials: JF
authors.lastname: Glatz
mesh_terms: D002352:Carrier Proteins / Q000302:isolation & purification*; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D005227:Fatty Acids / Q000378:metabolism*; D006801:Humans; D007525:Isoelectric Focusing; D009206:Myocardium / Q000737:chemistry*; D009363:Neoplasm Proteins; D015203:Reproducibility of Results; D013056:Spectrophotometry, Ultraviolet; D025521:Tumor Suppressor Proteins
chemical_list: D002352:Carrier Proteins; C107608:FABP7 protein, human; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D005227:Fatty Acids; D009363:Neoplasm Proteins; D025521:Tumor Suppressor Proteins