NR ARRB
AU Kuczius,T.; Buschmann,A.; Zhang,W.; Karch,H.; Becker,K.; Peters,G.; Groschup,M.H.
TI Cellular prion protein acquires resistance to proteolytic degradation following copper ion binding
QU Biological Chemistry 2004 Aug; 385(8): 739-47
PT journal article
AB The conversion of cellular prion protein (PrPc) into its pathological isoform (PrPsc) conveys an increase in hydrophobicity and induces a partial resistance to proteinase K (PK). Interestingly, co-incubation with high copper ion concentrations also modifies the solubility of PrPc and induces a partial PK resistance which was reminiscent of PrPsc. However, concerns were raised whether this effect was not due to a copper-induced inhibition of the PK itself. We have therefore analyzed the kinetics of the formation of PK-resistant PrPc and excluded possible interference effects by removing unbound copper ions prior to the addition of PK by methanol precipitation or immobilization of PrPc followed by washing steps. We found that preincubation of PrPc with copper ions at concentrations as low as 50 microM indeed rendered these proteins completely PK resistant, while control substrates were proteolyzed. No other divalent cations induced a similar effect. However, in addition to this specific stabilizing effect on PrPc, higher copper ion concentrations in solution (>200 microM) directly blocked the enzymatic activity of PK, possibly by replacing the Ca2+ ions in the active center of the enzyme. Therefore, as a result of this inhibition the proteolytic degradation of PrPc as well as PrPsc molecules was suppressed.
MH Animals; Blotting, Western; Calcium/chemistry; Cations/chemistry; Cattle; Colorimetry; Copper/antagonists & inhibitors/*chemistry; Edetic Acid/pharmacology; Endopeptidase K/*chemistry; Enzyme-Linked Immunosorbent Assay; Prions/*chemistry/isolation & purification; Protein Binding; Research Support, Non-U.S. Gov't; Sheep
AD Institute for Hygiene, University Hospital Münster, Robert-Koch-Str. 41, D-48149 Münster, Germany.
SP englisch
PO Deutschland