GPI-anchorless human prion protein is secreted and glycosylated but lacks superoxide dismutase activity.

نویسندگان

  • Akikazu Sakudo
  • Izuru Nakamura
  • Shotaro Tsuji
  • Kazuyoshi Ikuta
چکیده

Prion protein (PrP) is a glycosylphosphatidylinositol (GPI)-anchored membrane protein that is thought to play a role in anti-oxidative stress. It remains controversial whether PrP elicits superoxide dismutase (SOD) activity itself or indirectly by activating cellular SOD. Our previous studies showed that soluble PrP produced by a baculovirus expression system did not exhibit any SOD activity in a marginally glycosylated form. In the present study, we developed a mammalian expression system for a truncated soluble form of human prion protein with the native signal peptide but without a GPI-anchor site, driven by the peptide chain elongation factor 1alpha promoter in stably transfected rabbit-kidney epithelial RK13 cells, to investigate the SOD activity of mammalian PrP. This recombinant product, denoted sPrP, is secreted in large quantities in medium and can be isolated in very high purity and yield (more than 1 mg sPrP per 2 litres medium). Characterization by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and tunicamycin treatment revealed that a fully glycosylated form of sPrP was secreted from the cells. SOD activity in cell lysate showed a decrease in sPrP-expressing RK13 cells and an increase in wild-type PrP-expressing RK13 cells compared to empty vector-transfected RK13 cells or parent cells. Purified or immunoprecipitated sPrP did not show any SOD activity. In conclusion, the GPI-anchor site, but not glycosylation, appears to be essential for the secretion of PrP. In addition, mammalian PrP itself does not act as a functional SOD enzyme.

منابع مشابه

Characterization of the properties and trafficking of an anchorless form of the prion protein.

Conversion of PrP(C) into PrP(Sc) is the central event in the pathogenesis of transmissible prion diseases. Although the molecular basis of this event and the intracellular compartment where it occurs are not yet understood, the association of PrP with cellular membranes and in particular its presence in detergent-resistant microdomains appears to be of critical importance. In addition it appea...

متن کامل

Post-translational modifications in PrP expand the conformational diversity of prions in vivo

Misfolded prion protein aggregates (PrPSc) show remarkable structural diversity and are associated with highly variable disease phenotypes. Similarly, other proteins, including amyloid-β, tau, α-synuclein, and serum amyloid A, misfold into distinct conformers linked to different clinical diseases through poorly understood mechanisms. Here we use mice expressing glycophosphatidylinositol (GPI)-a...

متن کامل

Cells expressing anchorless prion protein are resistant to scrapie infection.

The hallmark of transmissible spongiform encephalopathies (TSEs or prion diseases) is the accumulation of an abnormally folded, partially protease-resistant form (PrP-res) of the normal protease-sensitive prion protein (PrP-sen). PrP-sen is attached to the cell membrane by a glycosylphosphatidylinositol (GPI) anchor. In vitro, the anchor and the local membrane environment are important for the ...

متن کامل

Defining the Conformational Features of Anchorless, Poorly Neuroinvasive Prions

Infectious prions cause diverse clinical signs and form an extraordinary range of structures, from amorphous aggregates to fibrils. How the conformation of a prion dictates the disease phenotype remains unclear. Mice expressing GPI-anchorless or GPI-anchored prion protein exposed to the same infectious prion develop fibrillar or nonfibrillar aggregates, respectively, and show a striking diverge...

متن کامل

Detection of the GPI-anchorless prion protein fragment PrP226* in human brain

BACKGROUND The accumulation of the misfolded forms of cellular prion protein, i.e. prions (PrPSc), in the brain is one of the crucial characteristics of fatal neurodegenerative disorders, called transmissible spongiform encephalopathies (TSEs). Cellular prion protein is normally linked to the cell surface by the glycosylphosphatidylinositol (GPI) anchor. There is accumulating evidence that the ...

متن کامل

ذخیره در منابع من


  با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید

متن کامل
عنوان ژورنال:
  • International journal of molecular medicine

دوره 21 2  شماره 

صفحات  -

تاریخ انتشار 2008