Iron(III)-phosphoprotein chelates: stoichiometric equilibrium constant for interaction of iron(III) and phosphorylserine residues of phosvitin and casein
Estimates of the strength of iron binding to model phosphoproteins were obtained from equilibrium dialysis experiments. Iron-free phosvitin (chicken and frog) or alpha sl-casein (cow) was dialyzed against the iron(III) chelates of nitrilotriacetate (NTA), )ethylenedinitrilo)tetraacetate (EDTA), or citrate. Protein-bound metal was measured at equilibrium; competition of chelator and phosphoprotein for iron(III) was determined by reference to comprehensive equilibrium equations presented in the Appendix. Analysis of the iron-binding data for phosvitin suggested that clusters of di-O-phosphorylserine residues (SerP.SerP) were the most probable iron-binding sites. A stoichiometric equilibrium constant of 10(18.0) was calculated for the formation of the Fe3+(SerP.SerP) chelate. When comared on the basis of phosphate content, casein bound iron more weakly than phosvitin. However, if the stoichiometric equilibrium constant for the formation of the casein Fe3+(SerP.SerP) chelate (10(17.5) was adjusted to account for the fact that a smaller percentage of casein phosphoserines occurs in di-O-phosphorylserine clusters, the affinity of casein and phosvitin for iron was very similar. A theoretical comparison showed that the "strengths" of the ferric chelates can be ranked: EDTA greater than phosphoprotein di-O-phosphorylserine greater than citrate greater than NTA.