In mammals, the
iron storage and detoxification
protein ferritin is composed of two functionally and genetically distinct subunit types, H (heavy) and L (light). The two subunits co-assemble in various ratios, with a tissue specific distribution, to form shell-like
protein structures of 24 subunits within which a mineralized
iron core is stored. The H-subunits possess
ferroxidase centers that catalyze the rapid oxidation of ferrous
ions, whereas the L-subunit does not have such centers and is believed to play an important role in electron transfer reactions that occur during the uptake and release of
iron. Pathogenic mutations on the L-chain lead to
neuroferritinopathy, a
neurodegenerative disease characterized by abnormal accumulation of
ferritin inclusion bodies and
iron in the central nervous system. Here, we have characterized the thermal stability,
iron loading capacity,
iron uptake, and
iron release properties of
ferritin heteropolymers carrying the three pathogenic
L-ferritin mutants (L154fs, L167fs, and L148fs, which for simplicity we named Ln1, Ln2 and Ln3, respectively), and a non-pathogenic variant (L135P) bearing a single substitution on the 3-fold axes of L-subunits. The UV-Vis data show a similar
iron loading capacity (ranging between 1800 to 2400 Fe(iii)/shell) for all
ferritin samples examined in this study, with Ln2 holding the least amount of
iron (i.e. 1800 Fe(iii)/shell). The three pathogenic
L-ferritin mutants revealed higher rates of
iron oxidation and
iron release, suggesting that a few mutated L-chains on the heteropolymer have a significant effect on
iron permeability through the
ferritin shell. DSC thermograms showed a strong destabilization effect, the severity of which depends on the location of the frameshift mutations (i.e. wt heteropolymer
ferritin ≅ homopolymer H-chain > L135P > Ln2 > Ln1 > Ln3). Variant L135P had only minor effects on the
protein functionality and stability, suggesting that local melting of the 3-fold axes in this variant may not be responsible for
neuroferritinopathy-like disorders. The data support the hypothesis that hereditary neuroferritinopathies are due to alterations of
ferritin functionality and lower physical stability which correlate with the frameshifts introduced at the C-terminal sequence and explain the dominant transmission of the disorder.