Characterization of the transition-metal-binding properties of hepcidin.
Accumulating evidence suggests that hepcidin, a 25-residue peptide hormone, is the master regulator of iron metabolism. Further evidence suggests that the five N-terminal amino acids are crucial for mediating its biological function. With a histidine residue at position 3, this region also has the potential to bind bivalent metal ions. To characterize this hepcidin-metal interaction in detail, the present study utilizes electrospray MS to measure the binding of a range of metal ions to wild-type and mutant human and murine hepcidins. In addition, the biological effects of these point mutations were tested on Caco-2 and HEK-293T human cell lines and in mice. Our results show that hepcidin-25 can form complexes with copper, nickel and zinc; however, we failed to detect any hepcidin-25 binding to either ferric or ferrous ions. The greatest affinity observed was between hepcidin-25 and copper with a dissociation constant <<1 microM. Substituting the histidine residue at position 3 in human hepcidin-25 and comparably the asparagine residue at position 3 in murine hepcidin-25 with an alanine residue markedly diminished the affinity for copper. The amino acid substitutions also decreased the biological activity of hepcidin-25; namely repression of ferroportin protein levels and hypoferraemia. In summary, the high affinity of hepcidin for copper suggests that hepcidin could bind copper in vivo and this may be of biological relevance.
Tselepis
Chris
C
University of Birmingham, UK. c.tselepis@bham.ac.uk
Ford
Samuel J
SJ
McKie
Andrew T
AT
Vogel
Wolfgang
W
Zoller
Heinz
H
Simpson
Robert J
RJ
Diaz Castro
Javier
J
Iqbal
Tariq H
TH
Ward
Douglas G
DG
D019943:Amino Acid Substitution; D000818:Animals; D023181:Antimicrobial Cationic Peptides / Q000235:genetics / Q000378:metabolism*; D002413:Cations, Divalent / Q000378:metabolism; D002460:Cell Line; D003300:Copper / Q000378:metabolism*; D064451:Hepcidins; D006801:Humans; D019216:Metals, Heavy / Q000378:metabolism; D051379:Mice; D017354:Point Mutation; D011485:Protein Binding; D021241:Spectrometry, Mass, Electrospray Ionization; D028561:Transition Elements / Q000378:metabolism*
D023181:Antimicrobial Cationic Peptides; D002413:Cations, Divalent; C578702:HAMP protein, human; C579599:Hamp protein, mouse; D064451:Hepcidins; D019216:Metals, Heavy; D028561:Transition Elements; D003300:Copper