Date of Award

Winter 1986

Project Type

Dissertation

Program or Major

Chemistry

Degree Name

Doctor of Philosophy

Abstract

Kinetic studies of the uptake of iron by transferrin from iron-pyrophosphate indicate that the reaction proceeds via an intermediate mixed ligand complex of the type PP(,i)-Fe-transferrin-CO(,3)('2-). The EPR spectrum of the "intermediate" complex was determined by computer modelling of the kinetic data and deconvolution of the experimental spectra. The distinct spectral characteristics of the intermediate indicate that the coordination environment of the iron is different from that of either of the reactants, consistent with formation of a complex in which iron is coordinated to both transferrin and pyrophosphate.

Studies of the interaction of sodium cyanide with iron(III)-transferrin demonstrate that a low-spin cyanide adduct of transferrin is produced which exhibits a rhombic EPR spectrum with principal components at g = 1.92, 2.15 and 2.34. The stoichiometry of adduct formation is: Fe(,c)-Tf-HCO(,3) + 2 CN('-) (--->) Fe(,c)-Tf-(CN)(,3) + HCO(,3)('-) where adduct formation only occurs at the C-terminal domain and the subscript "c" designates this half of the protein. This is the first example of an inorganic anion substituting for bicarbonate in transferrin. The results also show that ligands in the first coordination sphere of iron bound at the specific iron binding site in transferrin are susceptible to displacement by chelators in solution.

Nitrate binding to transferrin was studied using N-15 paramagnetic relaxation enhancement. Significant relaxation enhancement of the N-15 nucleus in the presence of bound paramagnetic iron was observed, indicating that the anion binding site is close to the metal on the protein. The metal anion distance was estimated to be in the range of eight to fifteen Angstroms.

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