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. 1991 Jul 15;88(14):6068–6071. doi: 10.1073/pnas.88.14.6068

Crosslinking of hemin to a specific site on the 90-kDa ferritin repressor protein.

J J Lin 1, M M Patino 1, L Gaffield 1, W E Walden 1, A Smith 1, R E Thach 1
PMCID: PMC52023  PMID: 2068086

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.

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