Copper(II) interaction with prion peptide fragments encompassing histidine residues within and outside the octarepeat domain: speciation, stability constants and binding details.
A 31-mer polypeptide, which encompasses residues 84-114 of human prion protein HuPrP(84-114) and contains three histidyl residues, namely one from the octarepeat (His85) and two histidyl residues from outside the octarepeat region (His96 and His111), and its mutants with two histidyl residues HuPrP(84-114)His85Ala, HuPrP(84-114) His96Ala, HuPrP(84-114)His111Ala and HuPrP(91-115) have been synthesised and their Cu2+ complexes studied by potentiometric and spectroscopic (UV/Vis, CD, EPR, ESI-MS) techniques. The results revealed a high Cu2+-binding affinity of all peptides, and the spectroscopic studies made it possible to clarify the coordination mode of the peptides in the different complex species. The imidazole nitrogen donor atoms of histidyl residues are the exclusive metal-binding sites below pH 5.5, and they have a preference for macrochelate structure formation. The deprotonation and metal-ion coordination of amide functions take place by increasing the pH; all of the histidines can be considered to be independent metal-binding sites in these species. As a consequence, di- and trinuclear complexes can be present even in equimolar samples of the metal ion and peptides, but the ratios of polynuclear species do not exceed the statistically expected ones; this excludes the possibility of cooperative Cu2+ binding. The species with a (N(im),N,N)-binding mode are favoured around pH 7, and their stability is enhanced by the macrochelation from another histidyl residue in the mononuclear complexes. The independence of the histidyl sites results in the existence of coordination isomers and the preference for metal binding follows the order of: His111>His96>His85. Deprotonation and metal-ion coordination of the third amide functions were detected in slightly alkaline solutions at each of the metal-binding sites; all had a (N(im),N,N,N)-coordination mode. Spectroscopic measurements also made it clear that the four lysyl amino groups of the peptides are not metal-binding sites in any cases.
Osz
Katalin
K
Department of Inorganic and Analytical Chemistry, University of Debrecen, 4010 Debrecen, Hungary.
Nagy
Zoltán
Z
Pappalardo
Giuseppe
G
Di Natale
Giuseppe
G
Sanna
Daniele
D
Micera
Giovanni
G
Rizzarelli
Enrico
E
Sóvágó
Imre
I
D002942:Circular Dichroism; D003300:Copper / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy; D006639:Histidine / Q000737:chemistry*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry*; D011199:Potentiometry; D011328:Prions / Q000737:chemistry*; D011968:Receptors, Histamine / Q000737:chemistry
D010446:Peptide Fragments; D011328:Prions; D011968:Receptors, Histamine; D006639:Histidine; D003300:Copper