Intact human ceruloplasmin is required for the incorporation of iron into human ferritin.
We have previously reported several studies on the loading of iron into ferritin by ceruloplasmin using proteins from rats. Loading iron into human ferritin using human serum ceruloplasmin is complicated by the fact that human ceruloplasmin is very susceptible to proteolysis (T. P. Ryan, T. A. Grover, and S. D. Aust, 1992, Arch. Biochem Biophys. 293, 1-8). The present study investigated the effect of proteolysis on the ability of human ceruloplasmin to load iron into human ferritin. SDS-PAGE revealed one major band with an apparent molecular weight of 116 kDa for a proteolytically degraded form of ceruloplasmin versus a 132-kDa band for an intact form of the enzyme. Both forms of the enzyme possessed ferroxidase activity, although that of the proteolytically degraded enzyme was approximately twofold less than that of the intact enzyme (4.9 nmol (min)-1 vs 8.3 nmol (min)-1). Only the intact form of ceruloplasmin was able to catalyze iron loading into ferritin without altering the physical characteristics of the ferritin protein during the process. Abnormal migration in nondenaturing PAGE gels, as well as a decrease in the amount of detectable ferritin protein, was observed when ferritin was incubated with iron alone or with proteolytically degraded ceruloplasmin and iron. It was concluded that the structural integrity of ceruloplasmin is required for the enzyme to effectively catalyze iron loading into ferritin.
Van Eden
M E
ME
Biotechnology Center, Utah State University, Logan 84322-4705, USA.
Aust
S D
SD
D000818:Animals; D002384:Catalysis; D002570:Ceruloplasmin / Q000737:chemistry / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D005293:Ferritins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D007501:Iron / Q000378:metabolism*; D008970:Molecular Weight; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D010446:Peptide Fragments; D011994:Recombinant Proteins; D005293:Ferritins; D007501:Iron; D002570:Ceruloplasmin