Functional and immunological analysis of recombinant mouse H- and L-ferritins from Escherichia coli.

The production and characterization of recombinant mouse H- and L-ferritin chains from Escherichia coli are described. The proteins were efficiently expressed and purified with yields of 7-40 mg per liter of cell culture. They had the expected molecular mass and showed a physical stability analogous to that of the corresponding human ferritins. Mouse H- and L-ferritins had a very similar mobility on denaturing SDS-PAGE, but could be readily separated on nondenaturing PAGE because of the distinct slow mobility of mouse L-ferritin. Direct comparative experiments showed that mouse and human H-ferritins had the same iron incorporation activity, whereas mouse L-ferritin incorporated iron less efficiently than human L-ferritin. The difference was attributed to the substitution of a residue exposed on the cavity surface (Glu140 --> Lys) in mouse L-ferritin, a hypothesis confirmed by the finding that the mouse L-ferritin mutant Lys140-Glu incorporated iron as efficiently as human L-ferritin. Rabbit antisera elicited by the recombinant mouse ferritins were specific for the H- and L-chains and did not cross-react with the human ferritins. The antibodies and the derived specific ELISA assays allow the determination of H- and L-ferritins in mouse tissues.