The catalytic activity of horse spleen apoferritin. Preliminary kinetic studies and the effect of chemical modification.
1. Horse spleen apoferritin catalyses the oxidation of Fe(2+) to Fe(3+) with molecular O(2) as electron acceptor under conditions where a number of other proteins have no such effect. The product is similar to ferritin by a number of criteria. 2. The progress curve is hyperbolic and the increase in initial velocity is linear with increasing apoferritin concentration. With respect to Fe(2+) the reaction follows Michaelis-Menten kinetics. The pH-dependence of the reaction was determined between pH4.3 and 6.0. 3. Modification of both tryptophan residues/apoferritin subunit with 2-nitrophenylsulphenyl chloride does not affect either k(cat.) or K(m) for the oxidation. Neither does the guanidination of seven out of nine lysine residues/subunit, the modification of nine out of ten arginine residues/subunit with cyclohexanedione, or the nitration of one out of five tyrosine residues/subunit with tetranitromethane. 4. The carboxymethylation of two out of three cysteine residues/subunit and of one out of six histidine residues/subunit can be achieved with iodoacetic acid. This carboxymethylated apoferritin is completely inactive in Fe(2+) oxidation. 5. Apoferritin does not take up Fe(3+). It appears from these results that Fe(2+) is the form in which iron is taken up by ferritin in a reaction where the protein acts as an enzyme which traps the product in the interior of the protein shell.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001059:Apoproteins; D002247:Carbon Isotopes; D002499:Centrifugation, Density Gradient; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D005293:Ferritins / Q000032:analysis / Q000378:metabolism*; D006146:Guanidines; D006736:Horses; D006863:Hydrogen-Ion Concentration; D007461:Iodoacetates; D007501:Iron / Q000032:analysis; D007700:Kinetics; D008697:Methane; D008854:Microscopy, Electron; D009574:Nitro Compounds; D009578:Nitrobenzenes; D010100:Oxygen; D013053:Spectrophotometry; D013056:Spectrophotometry, Ultraviolet; D013154:Spleen / Q000201:enzymology*; D013434:Sulfenic Acids; D013997:Time Factors; D014461:Ultracentrifugation
D000596:Amino Acids; D001059:Apoproteins; D002247:Carbon Isotopes; D006146:Guanidines; D007461:Iodoacetates; D009574:Nitro Compounds; D009578:Nitrobenzenes; D013434:Sulfenic Acids; D005293:Ferritins; D007501:Iron; D008697:Methane; D010100:Oxygen