title: Crosslinking of hemin to a specific site on the 90-kDa ferritin repressor protein.
abstract: Incubation of a 90-kDa ferritin repressor protein (FRP) with small amounts of radiolabeled hemin resulted in the formation of a strong interaction between the two that was stable to SDS/PAGE. (We refer to this interaction as a "crosslink," without intending to imply knowledge as to its chemical nature.) Of seven other proteins tested individually, only apohemopexin and bovine serum albumin showed similar crosslinking ability, albeit to a much lower extent. [14C]Hemin specifically crosslinked to FRP in the presence of a 50-fold excess of total wheat germ proteins. Inclusion of catalase did not prevent the reaction of hemin with FRP, suggesting that H2O2 is not involved. The subsequent addition of a stoichiometric amount of apohemopexin did not reverse the reaction. Exhaustive digestion of the complex with Staphylococcus aureus V8 protease produced a major labeled peptide of 17 kDa. These results show the existence of a highly specific, uniquely reactive hemin binding site on FRP.
authors.affiliation: Department of Biology, Washington University, St. Louis, MO 63130.
authors.forename: J J
authors.initials: JJ
authors.lastname: Lin
authors.forename: M M
authors.initials: MM
authors.lastname: Patino
authors.forename: L
authors.initials: L
authors.lastname: Gaffield
authors.forename: W E
authors.initials: WE
authors.lastname: Walden
authors.forename: A
authors.initials: A
authors.lastname: Smith
authors.forename: R E
authors.initials: RE
authors.lastname: Thach
mesh_terms: D001665:Binding Sites; D002352:Carrier Proteins / Q000302:isolation & purification / Q000378:metabolism*; D003432:Cross-Linking Reagents; D005293:Ferritins / Q000378:metabolism; D006427:Hemin / Q000378:metabolism*; D035941:Iron Regulatory Protein 1; D035925:Iron-Regulatory Proteins; D007700:Kinetics; D008970:Molecular Weight
chemical_list: D002352:Carrier Proteins; D003432:Cross-Linking Reagents; D035925:Iron-Regulatory Proteins; D006427:Hemin; D005293:Ferritins; D035941:Iron Regulatory Protein 1