Identification, functional characterization, and cryo-EM structural analysis of novel ferritin subunits in Turbo...

Turbo sazae, an edible gastropod, accumulates high levels of iron in its digestive gland, and the molecular mechanism underlying this accumulation has remained elusive. This study identified the proteins responsible for the iron accumulation and characterized their function and structure. We isolated two novel ferritins, TsFerA and TsFerB, from the digestive gland using high-performance liquid chromatography and inductively coupled plasma mass spectrometry. Gene expression analyses revealed that both genes were specifically expressed in the digestive gland. Recombinant TsFerA (rTsFerA) possessed ferroxidase (EC1.16.3.1) activity, whereas recombinant TsFerB (rTsFerB) showed no such activity. This result indicated that rTsFerA functions as an H-chain-like subunit responsible for iron oxidation, while rTsFerB acts as an L-chain-like subunit involved in iron core nucleation. Furthermore, we determined the structures of rTsFerA and rTsFerB using cryo-electron microscopy, at a resolution of 2.19 and 2.17 Å, respectively. The protein structures revealed a conserved ferroxidase center in rTsFerA, whereas key catalytic residues were substituted in rTsFerB. These findings demonstrate that T. sazae utilizes a cooperative system composed of two functionally distinct ferritin subunits to efficiently and safely store iron. This work clarifies the molecular mechanisms of iron metabolism in marine organisms, particularly in gastropods, revealing an optimized strategy to cope with massive iron influx.