A spectroscopic and voltammetric study of the pH-dependent Cu(II) coordination to the peptide GGGTH: relevance to the fifth Cu(II) site in the prion protein.
The GGGTH sequence has been proposed to be the minimal sequence involved in the binding of a fifth Cu(II) ion in addition to the octarepeat region of the prion protein (PrP) which binds four Cu(II) ions. Coordination of Cu(II) by the N- and C-protected Ac-GGGTH-NH(2) pentapeptide (P(5)) was investigated by using potentiometric titration, electrospray ionization mass spectrometry, UV-vis spectroscopy, electron paramagnetic resonance (EPR) spectroscopy and cyclic voltammetry experiments. Four different Cu(II) complexes were identified and characterized as a function of pH. The Cu(II) binding mode switches from NO(3) to N(4) for pH values ranging from 6.0 to 10.0. Quasi-reversible reduction of the [Cu(II)(P(5))H(-2)] complex formed at pH 6.7 occurs at E (1/2)=0.04 V versus Ag/AgCl, whereas reversible oxidation of the [Cu(II)(P(5))H(-3)](-) complex formed at pH 10.0 occurs at E (1/2)=0.66 V versus Ag/AgCl. Comparison of our EPR data with those of the rSHaPrP(90-231) (Burns et al. in Biochemistry 42:6794-6803, 2003) strongly suggests an N(3)O binding mode at physiological pH for the fifth Cu(II) site in the protein.
Hureau
Christelle
C
Laboratoire de Géophysique Interne et Tectonophysique, Université Joseph Fourier, BP 53, 38041 Grenoble Cedex 9, France. christelle.hureau@cea.fr
Charlet
Laurent
L
Dorlet
Pierre
P
Gonnet
Florence
F
Spadini
Lorenzo
L
Anxolabéhère-Mallart
Elodie
E
Girerd
Jean-Jacques
JJ
D003300:Copper / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy; D006863:Hydrogen-Ion Concentration; D009842:Oligopeptides / Q000737:chemistry*; D011199:Potentiometry; D011328:Prions / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization; D013056:Spectrophotometry, Ultraviolet
D009842:Oligopeptides; D011328:Prions; D003300:Copper