Iron-Storage Mechanism of Ferritin

Iron-Storage Mechanism of Ferritin

Honarmand Ebrahimi, K.

Hagen, W. (promotor)

Applied Sciences

Biotechnology

2013-05-01

Storage of Fe(III) is a common mechanism by which the cellular machinery controls the availability of Fe(II) and Fe(III) for biosynthesis of iron-containing cofactors of enzymes which are involved in several essential biological processes, including oxidative phosphorylation. The conserved 24-meric iron-storage protein ferritin has been identified in many organisms to control the availability of Fe(II) by oxidizing the excess Fe(II) and storing the Fe(III) oxidation product in a soluble and nontoxic form. A conserved diiron center, the ferroxidase center, is responsible for catalytic oxidation of Fe(II), the ferroxidase reaction. The mechanism by which ferritin stores the Fe(III) is not fully understood, and the current models in the literature suggest different mechanisms for the functioning of ferritins from different Domains of life. Moreover, a structural gene for a 24-meric ferritin has not been found in some organisms including Pyrococcus abyssi or Pyrococcus horikoshii. Below we first outline methods which can be used to measure ferroxidase activity of different proteins. As an example we measure the ferroxidase activity of two proteins, human H ferritin and ceruloplasmin, and that of a synthetic peptide. Subsequently, using these techniques we study the mechanism of iron oxidation of a ferritin from hyperthermophilic archaeal anaerobe Pyrococcus furiosus. We then employ new experimental approaches using isotopically labeled 57Fe(II) to compare the iron-storage mechanism of P. furiosus ferritin with that of eukaryotic human H ferritin. We demonstrate that, conflicting with the current models in the literature these proteins employ a common mechanism to store the Fe(III) oxidation product. We suggest that this mechanism is general from archaea to eukaryotes. Finally, we carry out the in-vitro biochemical characterization of a new member of the ferritin superfamily of proteins that unlike the 24-meric ferritin is monomeric in the absence of iron. We name this protein archaeoferritin and we show that monomers oxidize Fe(II) and reversibly assemble to form Fe(III)-storing oligomeric structures comparable to that of ferritin.

Ferritin,
Iron-storage,
Ferroxidase,
Archaeoferritin,
amyloid precursor protein,
EPR spectroscopy,
Mossbauer spectroscopy,
PfFtn,
HuHF,
BfMF,
BfHF,
Eukaryotic ferritin,
Microbial ferritin,
EcFtnA,
Diiron catalytic center,
Alzheimer's disease,
Isothermal titration calorimetry,
Bullfrog Ferritin,
Bacterioferritin,
Dps protein,
Blue intermediate,
Peroxodiferric intermediate,
Tyrosine,
Frataxin,
MALDI-ToF,
Mass spectrometry,
Iron oxidation,
ferroxidase reaction,

https://doi.org/10.4233/uuid:dd409de5-dbbe-4124-9f94-9b67caef35cb

2013-05-02

9789088915994

Institutional Repository

doctoral thesis

(c) 2013 Honarmand Ebrahimi, K.