Kinetic analysis of bovine spleen apoferritin and recombinant H and L chain homopolymers: iron uptake suggests early stage H chain ferroxidase activity and second stage L chain cooperation.
Ferritin utilizes ferroxidase activity to incorporate iron. Iron uptake kinetics of bovine spleen apoferritin (H: L = 1 : 1.1) were compared with those of recombinant H chain ferritin and L chain ferritin homopolymers. H chain ferritin homopolymer showed an iron uptake rate identical to bovine spleen apoferritin (0.19 and 0.21 mmol/min/micromol of protein, respectively), and both showed iron concentration-dependent uptake. In contrast, the L chain homopolymer, which lacks ferroxidase, did not incorporate iron and showed the same level of iron autoxidation in the absence of ferritin. Bovine spleen apoferritin was shown to have two iron concentration-dependent uptake pathways over a range of 0.02-0.25 mM ferrous ammonium sulfate (FAS) by an Eadie-Scatchard plot (v/[FAS] versus v), whereas the H chain ferritin homopolymer was found to have only one pathway. Of the two Km values found in bovine spleen apoferritin, the lower mean Km value was 9.0 microM, while that of the H chain homopolymer was 11.0 microM. H chain ferritin homopolymer reached a saturating iron uptake rate at 0.1 mM FAS, while bovine spleen apoferritin incorporated more iron even at 0.25 mM FAS. These results suggest that the intrinsic ferroxidase of ferritin plays a significant role in iron uptake, and the L chain cooperates with the H chain to increase iron uptake.
Orino
Koichi
K
Laboratory of Biochemistry, School of Veterinary Medicine and Animal Sciences, Kitasato University, Aomori 034-8628, Japan. orino@vmas.kitasato-u.ac.jp
Harada
Satoshi
S
Natsuhori
Masahiro
M
Takehara
Kazuaki
K
Watanabe
Kiyotaka
K
D000818:Animals; D001052:Apoferritins / Q000378:metabolism*; D002417:Cattle; D002570:Ceruloplasmin / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D007501:Iron / Q000378:metabolism*; D007700:Kinetics; D013154:Spleen / Q000201:enzymology / Q000378:metabolism*; D013997:Time Factors
D001052:Apoferritins; D007501:Iron; D002570:Ceruloplasmin