The cellular prion protein binds copper in vivo

Abstract

The normal cellular form of prion protein (PrPC) is a precursor to the pathogenic protease-resistant forms (PrPSc) believed to cause scrapie, bovine spongiform encephalopathy (BSE) and Creutzfeldt-Jakob disease. Its amino terminus contains the octapeptide PHGGGWGQ, which is repeated four times and is among the best-preserved regions of mammalian PrPC. Here we show that the amino-terminal domain of PrPC exhibits five to six sites that bind copper (Cu(II)) presented as a glycine chelate. At neutral pH, binding occurs with positive cooperativity, with binding affinity compatible with estimates for extracellular, labile copper. Two lines of independently derived PrPC gene-ablated (Prnp0/0) mice exhibit severe reductions in the copper content of membrane-enriched brain extracts and similar reductions in synaptosomal and endosome-enriched subcellular fractions. Prnp0/0 mice also have altered cellular phenotypes, including a reduction in the activity of copper/zinc superoxide dismutase and altered electrophysiological responses in the presence of excess copper. These findings indicate that PrPC can exist in a Cu-metalloprotein form in vivo.

The Affinity of Copper Binding to the Prion Protein Octarepeat Domain: Evidence for Negative Cooperativity

“…For instance, equilibrium dialysis measurements performed on PrP(23-98) find a half-maximal binding at 5.9 μM and a Hill coefficient (n) of 3.4, indicating a very strong positive cooperativity through the micromolar range (8). Similar results were obtained from CD measurements of the d-d absorption band at 570 nm, arising from Cu 2+ binding to PrP (58-91) (25).…”

“…Clearly, the results reported here require not only confirmation in full-length PrP but also an assessment of the influence of non-octarepeat coordination sites (20,28) and other features that may affect Cu 2+ uptake. With regard to octarepeat coordination, however, our previously published experiments on full-length PrP Since the discovery of the physiological connection of PrP to copper, there have been intensive efforts to determine the precise Cu 2+ affinity (8,10,(21)(22)(23)(24)(25). As noted in the Introduction, there is wide disagreement with reported K d values distributed over nearly 8 orders of magnitude.…”

“…of PrP C is unknown, but recent investigations focus on the ability of the octarepeat domain to take up copper (5,(7)(8)(9)(10)(11). PrP C protects against apoptosis (12) and radical-mediated oxidative damage (13,14) and even stimulates nerve cell growth and development (15).…”

“…For an enzyme, one would expect high affinity, reflected through a low dissociation constant (K d ) (21), whereas buffering or sensing would require a lower affinity, with a K d near the concentration of extracellular copper (22). Unfortunately, investigations into the dissociation constant and the mechanism of copper uptake reveal highly disparate outcomes depending upon the measurement technique and the particular PrP construct (8,10,(21)(22)(23)(24)(25). Current estimates place K d between 10 −6 M (10,22,23) and 10 −14 M (21,24), an experimental uncertainty of 8 orders of magnitude.…”

Risikofaktoren der sporadischen Creutzfeldt-Jakob-Krankheit

“…Seine Funktion ist noch nicht geklärt. Zusammenhänge mit Kupferbindung und -transport, antioxidativen Wirkungen und zirkadianer Rhythmik werden derzeit noch diskutiert (Brown DR et al 1997a, Brown DR et al 1997b, Tobler et al 1996. Außerdem könnte das PrP C in der Neurotransmission an Synapsen eine essentielle Rolle spielen (Collinge et al 1994, Carleton et al 2001, Hu et al 2007).…”