Interaction of the cellular prion protein with raft-like lipid membranes

Abstract Conversion of the cellular isoform of the prion protein (PrPC) into the disease-associated isoform (PrPSc) plays a key role in the development of prion diseases. Within its cellular pathway, PrPC undergoes several posttranslational modifications, i.e., the attachment of two N-linked glycans and a glycosyl phosphatidyl inositol (GPI) anchor, by which it is linked to the plasma membrane on the exterior cell surface. To study the interaction of PrPC with model membranes, we purified posttranslationally modified PrPC from transgenic Chinese hamster ovary (CHO) cells. The mono-, di- and oligomeric states of PrPC free in solution were analyzed by analytical ultracentrifugation. The interaction of PrPC with model membranes was studied using both lipid vesicles in solution and lipid bilayers bound to a chip surface. The equilibrium and mechanism of PrPC association with the model membranes were analyzed by surface plasmon resonance. Depending on the degree of saturation of binding sites, the concentration of PrPC released from the membrane into aqueous solution was estimated at between 10-9 and 10-7 M. This corresponds to a free energy of the insertion reaction of -48 kJ/mol. Consequences for the conversion of PrPC to PrPSc are discussed.

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Both lysine-clusters of the NH2-terminal prion-protein fragment PrP23-110 are essential for t-PA mediated plasminogen activation

Thrombosis and Haemostasis ◽

10.1160/th03-06-0382 ◽

2004 ◽

Vol 91 (03) ◽

pp. 465-472 ◽

Cited By ~ 16

Author(s):

Guido Epple ◽

Kristina Langfeld ◽

Michael Baier ◽

Hermann-Georg Holzhütter ◽

Eckart Köttgen ◽

...

Keyword(s):

Prion Protein ◽

Plasmon Resonance ◽

Cellular Prion Protein ◽

Plasminogen Activation ◽

Protein Fragment ◽

Chip Surface ◽

Mutant Proteins ◽

Lysine Residues ◽

Binding Curves

SummaryWe have recently shown that the NH2-terminal fragment (PrP23-110) of the human cellular prion protein (PrPc) stimulates t-PA mediated plasminogen activation. PrP23-110 contains an N-terminal lysine cluster (LC1; K23, K24, K27) and a C-terminal one (LC2; K101, K104, K106, K110). To study their biological function we have substituted all lysine residues of each cluster by alanine and generated the recombinant PrP proteins PrP23110sLC1 and PrP23-110sLC2. The ability of the mutant proteins to stimulate plasminogen activation was assayed. We found that both lysine clusters are essential for t-PA mediated plasminogen activation. We further studied the binding of soluble PrP23110 to immobilized t-PA or plasminogen using surface plasmon resonance. The recorded binding curves could not be modeled by classical 1:1 binding kinetics suggesting oligomerisation of PrP23-110. Further plasmon resonance studies show that indeed PrP23-110 binds to itself and that glycosaminoglycans modify this interaction. Binding of t-PA or plasminogen to PrP23-110 was no longer influenced by glycosaminoglycans when PrP23-110 was immobilized on the chip surface. Thus a possible role of heparin as a cofactor in the stimulation of plasminogen activation by t-PA could be the generation of a PrP23-110 form with both lysine clusters accessible for binding of t-PA and plasminogen.

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Interfacial water at synthetic and natural lipid bilayers probed by vibrational sum-frequency generation spectroscopy

Biophysical Bulletin ◽

10.26565/2075-3810-2020-43-09 ◽

2020 ◽

Keyword(s):

Lipid Bilayers ◽

Pure Water ◽

Lipid Vesicles ◽

Sum Frequency Generation ◽

Model Membranes ◽

Average Charge ◽

Interfacial Water ◽

Sum Frequency ◽

Frequency Generation ◽

Natural Lipid

Background: At lipid interfaces, water plays a crucial role in carrying biological processes, so that there is a huge interest in unravelling the behaviour of water close to membranes. At charged bio-interfaces, water dipoles form an organized layer. Probing such an interfacial thin layer buried between macroscopic bulk environments is a real challenge. Vibrational sum frequency generation (SFG) spectroscopy is intrinsically specific to interfaces, and has already proven to be an ideal tool to investigate model membranes and their surrounding water. Objectives: The goal of this work is to measure the vibrational SFG response of interfacial water around different model membranes — from easiest synthetic lipids to more complex natural lipids, — in order to use it as diagnostic signal able to distinguish the lipid bilayer interface by its charge properties. Materials and methods: Lipid bilayers made either of synthetic or natural lipids (Avanti Polar Lipids) were physisorbed on CaF2 prisms (Crystran), by using the method of the spontaneous fusion of lipid vesicles, to form so called solid-supported lipid bilayers (SSLBs). The model membranes were investigated by SFG spectroscopy at the solid/water interface. Results: The SFG response was measured between 3600 cm-1 and 2800 cm-1, where OH stretching vibrations of water molecules show-up. The SFG intensity of the OH peak maximum at 3125 cm-1 was recorded during the adsorption of lipid vesicles on the surface, and provided knowledge of the changes of the charge properties of the interface due to the adsorption of the model membranes. The SFG signal indicated that the organization of water was larger at negatively charged than at positively lipid interfaces, and reached the highest value with natural E. сoli сardiolipin layers. Moreover, when the full composition of natural lipids was unknown, the behaviour of the SFG response enabled establishing the charge characteristics of the corresponding lipid interfaces. Conclusion: The SFG response of water enabled estimating average charge behaviour of synthetic and natural lipid bilayers in pure water, thus paving the way to use the SFG signal of water as new diagnostic tool to identify lipid interfaces.

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Expression of cellular prion protein on blood cells: Potential functions in cell physiology and pathophysiology of transmissible spongiform encephalopathy diseases

Transfusion Medicine Reviews ◽

10.1053/tm.2001.26957 ◽

2001 ◽

Vol 15 (4) ◽

pp. 268-281 ◽

Cited By ~ 5

Author(s):

Jaroslav G. Vostal ◽

Karel Holada ◽

Jan Simak

Keyword(s):

Prion Protein ◽

Blood Cells ◽

Transmissible Spongiform Encephalopathy ◽

Potential Functions ◽

Cellular Prion Protein ◽

Spongiform Encephalopathy ◽

Cell Physiology

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Differential expression of cellular Prion protein in neural development in Familial Alzheimer’s Disease

10.26226/morressier.5b31ec2b2afeeb001345a47e ◽

2018 ◽

Author(s):

Cleide Dos Santos Souza

Keyword(s):

Alzheimer’S Disease ◽

Alzheimer's Disease ◽

Differential Expression ◽

Prion Protein ◽

Neural Development ◽

Cellular Prion Protein ◽

Familial Alzheimer’S Disease ◽

Familial Alzheimer's Disease

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Faculty Opinions recommendation of Cellular prion protein mediates impairment of synaptic plasticity by amyloid-beta oligomers.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1158625.619545 ◽

2009 ◽

Author(s):

Gerald Zamponi

Keyword(s):

Synaptic Plasticity ◽

Prion Protein ◽

Amyloid Beta ◽

Cellular Prion Protein

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Pharmacological inactivation of the prion protein by targeting a folding intermediate

Communications Biology ◽

10.1038/s42003-020-01585-x ◽

2021 ◽

Vol 4 (1) ◽

Author(s):

Giovanni Spagnolli ◽

Tania Massignan ◽

Andrea Astolfi ◽

Silvia Biggi ◽

Marta Rigoli ◽

...

Keyword(s):

Prion Protein ◽

Prion Diseases ◽

Cellular Prion Protein ◽

Surface Glycoprotein ◽

Folding Intermediate ◽

Biological Regulation ◽

Cell Surface Glycoprotein ◽

Folding Intermediates ◽

Protein Levels ◽

Small Molecule Ligands

AbstractRecent computational advancements in the simulation of biochemical processes allow investigating the mechanisms involved in protein regulation with realistic physics-based models, at an atomistic level of resolution. These techniques allowed us to design a drug discovery approach, named Pharmacological Protein Inactivation by Folding Intermediate Targeting (PPI-FIT), based on the rationale of negatively regulating protein levels by targeting folding intermediates. Here, PPI-FIT was tested for the first time on the cellular prion protein (PrP), a cell surface glycoprotein playing a key role in fatal and transmissible neurodegenerative pathologies known as prion diseases. We predicted the all-atom structure of an intermediate appearing along the folding pathway of PrP and identified four different small molecule ligands for this conformer, all capable of selectively lowering the load of the protein by promoting its degradation. Our data support the notion that the level of target proteins could be modulated by acting on their folding pathways, implying a previously unappreciated role for folding intermediates in the biological regulation of protein expression.

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