Production and characterization of recombinant heteropolymers of human ferritin H and L chains.
Vertebrate ferritins are iron storage proteins composed by 24 subunits of one or more types. The recombinant homopolymers of human ferritin H- and L-type chains differ in iron uptake and in physical stability, but the properties of heteropolymers with various proportions of H- and L-type chains cannot be predicted. Present study shows that unfolded human ferritin H- and L- type chains renature under similar conditions to form homopolymers indistinguishable from the native ones and that, when mixed, the unfolded H and L chains renature to form heteropolymers with restricted heterogeneity and with the expected H:L ratios. Seven of these ferritins with different H:L ratios were analyzed; electrophoretic mobility, immunological reactivity, and stability to guanidine denaturation varied as predicted, based on the homopolymers. In contrast, the rate of iron uptake, monitored by the variation of absorbance at 310 nm, increased in the ferritins that ranged in H chain content from 0 to 35%; further increments in H chains had no additional effect. This finding indicates that, under the present conditions, only a limited number of H chains are needed for the maximum rate of ferritin iron uptake. Variations of L- and H-type chains in vivo may thus have biological relevance.
Department of Biological and Technological Research, Scientific Institute San Raffaele, Milano, Italy.
P
P
Santambrogio
S
S
Levi
A
A
Cozzi
E
E
Rovida
A
A
Albertini
P
P
Arosio
D002942:Circular Dichroism; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005293:Ferritins / Q000096:biosynthesis* / Q000737:chemistry* / Q000302:isolation & purification; D019791:Guanidine; D006146:Guanidines; D006801:Humans; D007501:Iron / Q000378:metabolism; D007525:Isoelectric Focusing; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification
D006146:Guanidines; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D005293:Ferritins; D007501:Iron; D019791:Guanidine