Recombinant human hepcidin expressed in Escherichia coli isolates as an iron containing protein.
Hepcidin is a small peptide that acts as a regulator of systemic iron homeostasis. To study some of its functional properties, a synthetic cDNA for the minimal, 20-amino-acid, form of human hepcidin was cloned into different constructs for expression in Escherichia coli. The fusion ferritin-hepcidin produced molecules retaining most of ferritin structural and functional properties, including ferroxidase and iron incorporation activities. However, it showed spectroscopic properties compatible with the presence of iron-sulfur complexes on the hepcidin moiety, which was buried into protein cavity. Similar complexes were reconstituted by in vitro incubation of the iron-free protein with iron and sulfide salts. Two other unrelated fusion products were constructed, which, when expressed in E. coli, formed insoluble aggregates retaining a large proportion of total bacterial iron. Analysis of the solubilized preparations showed them to contain iron-sulfur complexes. We concluded that the cysteine-rich hepcidin acts as an iron-sequestering molecule during expression in E. coli. This may have implications for the biological functions of this key protein of iron metabolism.
Gerardi
Gianmario
G
Dipartimento Materno Infantile e Tecnologie Biomediche, Università di Brescia, A.O. Spedali Civili, Viale Europa 11, 25125 Brescia, Italy.
Biasiotto
Giorgio
G
Santambrogio
Paolo
P
Zanella
Isabella
I
Ingrassia
Rosaria
R
Corrado
Marcella
M
Cavadini
Patrizia
P
Derosas
Manuela
M
Levi
Sonia
S
Arosio
Paolo
P
D023181:Antimicrobial Cationic Peptides / Q000235:genetics* / Q000302:isolation & purification / Q000502:physiology*; D003001:Cloning, Molecular / Q000379:methods; D004926:Escherichia coli / Q000235:genetics; D005293:Ferritins / Q000235:genetics; D064451:Hepcidins; D006801:Humans; D007501:Iron / Q000378:metabolism*; D019159:Nonheme Iron Proteins / Q000235:genetics / Q000502:physiology*; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D012995:Solubility
D023181:Antimicrobial Cationic Peptides; C578702:HAMP protein, human; D064451:Hepcidins; D019159:Nonheme Iron Proteins; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D005293:Ferritins; D007501:Iron