Prion Protein Misfolding Structure Determination For PRPC Biology Essay

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The molecular structure of PrPC at atomic resolution has been determined by nuclear magnetic resonance(NMR) spectroscopy and X-ray crystallography. PrPC consists of a long and flexible amino terminal region spanning up to amino acid (aa) residue 121 and a structured carboxy terminal domain. This globular domain harbours two short sheet-forming anti-parallel beta-strands (aa 128 to 130 and aa 160 to 162 in murine PrPC) and three 􀀁-helices (helix I: aa 143 to 153; helix II: aa 171 to 192; helix III: aa 199 to 226 in murine PrPC) [28]. The length of the unprocessed translation product is 256 amino acids. In the course of its transit through the ER and Golgi apparatus, post-translational modifications occur, such as the removal of a N terminal signal sequence (1-22); the formation of an internal helix II and III stabilizing disulfide bond (between aa 179 and aa 214); the attachment of Nlinked oligosaccharide chains (at aa 180 and aa 196); and the replacement of the carboxy terminus (at aa 231) by a glycosylphosphatidylinositol (GPI) anchor. Fully processed (murine) PrPC therefore contains only 209 amino acids, representing codons 23-231 ofthe prion ORF.

However, on the basis of cryoelectron microscopy and by the means of structural modeling based on similar common protein structures, it has been discovered that PrPSc contains ß-sheets in the region of aa 81-95 to aa 171, while the carboxy terminal structure is supposedly preserved. These ß-sheets form a left-handed beta-helix. Three PrPSc molecules are believed to form a primary unit and therefore build the basis for the so-called scrapie-associated fibrils.

The increase in the content of beta -sheet structures results in insolubility in mild detergent fluids and causes partial resistance to enzymatic degradation of the pathogenic isoform PrPSc . If PrPSc is treated with proteolytic enzymes, only the N-terminal amino acids (aa) up to residues 81-95 (depending on the TSE agent and the proteolytic conditions) are digested [32, 33], leaving the remaining PrPSc reaching from aa 81-95 to aa 231. The increased resistance to proteolysis leads to an accumulation of PrPSc..

PrPSc particles which consist of only 14-28 PrP molecules exhibit the highest rate of infectivity and conversion. Protein folding and hence misfolding is determined by the primary structure of a polypeptide chain, but the complex process of protein folding kinetics has been a major topic for decades and is still not completely understood. Despite numerous models for protein folding, there also exist various theories as to how misfolding could be explained.

rapid refolding under physiological conditions has been shown for spider-silk proteins that form beta -sheet rich fibres contingent upon the rapid decrease of sodium, increase of potassium concentration and a drop in the pH

Misfolding can only take place when the native structure of a globular protein is at least partially unfolded or degraded. Spontaneous protein misfolding may occur more frequently under physiological conditions than is generally assumed. Cellular factors and pathways could be of major relevance in regards to disease prevention or initiation.

Chaperones may have a key role in preventing pathogenic effects of misfolding and aggregation.. As for PrP, chaperones have been shown to play an interchangeable role: certain heat shock proteins are able to promote conversion, whereas others inhibit misfolding . The chaperon BiP, which is present in the endoplasmatic reticulum (ER), has been shown to bind to certain forms of PrP that were retained in the ER due to incomplete processing . Within the ER, BiP is believed to maintain proper folding of PrP by binding to defective forms for an extended period of time. In this way, the defective forms can finally be degraded by the proteasomal pathway. Due to their occurrence in amyloids, there is evidence for the assumption that nucleic acids, lipids and glycosaminoglycans (GAGs) might play a role as cofactors in amyloidogenesis. For this reason they could be a useful therapeutic target not only for prion disorders but also for other protein misfolding diseases.

As common components of amyloid, GAGs are found in PrPSc in vivo, it has been shown that they facilitate the conversion of PrPC into PrPSc in vitro, as well as PrP-aggregation. Lipids and nucleic acids also bind to PrPC and are detectable in PrPSc -aggregates ; additionally, they may facilitate PrP-conversion by functioning as a scaffold that binds and concentrates PrPC in order to provide high amounts of substrate for a conversion into PrPSc .

Basically, two types of PrP conversion can be distinguished - induced misfolding and spontaneous, or non-seeded, PrPSc or PrPres forming. The latter is seen in inherited human TSEs,whereas the induced misfolding needs an infection to begin, e.g. through the oral intake of infected tissues.

It has been demonstrated by NMR spectroscopy that some disease related mutations of the human PrPC are located in a part of the protein that is involved in the maintenance of the hydrophobic core in the fibril . Amino acid mutations therefore do not necessarily alter the stability of PrP but might have some local effects on the protein interactions which are required for oligomerization into fibrillar species. The exposure of hydrophobic regions in intermediate states during protein folding could increase the tendency towards aggregation, and subsequently initiate - at a certain stage - the misfolding cascade, which ultimately leads to disease. Hydrophobic interactions play a crucial role in the formation of beta -sheets, as they bring fragments of a polypeptide chain in close proximity to each other.

The disease-promoting mutations in the human PrPC had a statistically significant tendency towards increasing local hydrophobicity with a possible change in interactions between PrP molecules and/or between PrP and hypothetical cofactors that might initiate subsequent fibril formation.

a peptide spanning aa residues 106-126 which displayed a neurotoxic effect on rat hippocampal neurons in vitro. Neurotoxicity could also be demonstrated in vivo . This part of the prion protein is therefore commonly referred to as neurotoxic peptide. These toxic characteristic are restricted exclusively to cells which express PrPC.

Because the prion fragment 106- 126 shares many properties of PrPSc , e.g. the ability to form beta-structures and partially protease-resistant fibrils, it is seen as a model for molecular mechanisms in neurodegeneration caused by prion protein misfolding.

The highly amyloigenic and hydrophobic palindrome AGAAAAGA is located between aa residue 113 and 120 of PrP 106-126. It is described as putative aggregation site [106], although this sequence requires its flanking parts to form fibrillar aggregates