Rabbit Prion Protein Research Articles

Y225A induces long-range conformational changes in human prion protein that are protective in Drosophila

Prion protein (PrP) misfolding is the key trigger in the devastating prion diseases. Yet the sequence and structural determinants of PrP conformation and toxicity are not known in detail. Here, we describe the impact of replacing Y225 in human PrP with A225 from rabbit PrP, an animal highly resistant to prion diseases. We first examined human PrP-Y225A by molecular dynamics simulations. We next introduced human PrP in Drosophila and compared the toxicity of human PrP-WT and Y225A in the eye and in brain neurons. Y225A stabilizes the β2-α2 loop into a 310-helix from six different conformations identified in WT and lowers hydrophobic exposure. Transgenic flies expressing PrP-Y225A exhibit less toxicity in the eye and in brain neurons and less accumulation of insoluble PrP. Overall, we determined that Y225A lowers toxicity in Drosophila assays by promoting a structured loop conformation that increases the stability of the globular domain. These findings are significant because they shed light on the key role of distal α-helix 3 on the dynamics of the loop and the entire globular domain.

Insight From Animals Resistant to Prion Diseases: Deciphering the Genotype - Morphotype - Phenotype Code for the Prion Protein.

Prion diseases are a group of neurodegenerative diseases endemic in humans and several ruminants caused by the misfolding of native prion protein (PrP) into pathological conformations. Experimental work and the mad-cow epidemic of the 1980s exposed a wide spectrum of animal susceptibility to prion diseases, including a few highly resistant animals: horses, rabbits, pigs, and dogs/canids. The variable susceptibility to disease offers a unique opportunity to uncover the mechanisms governing PrP misfolding, neurotoxicity, and transmission. Previous work indicates that PrP-intrinsic differences (sequence) are the main contributors to disease susceptibility. Several residues have been cited as critical for encoding PrP conformational stability in prion-resistant animals, including D/E159 in dog, S167 in horse, and S174 in rabbit and pig PrP (all according to human numbering). These amino acids alter PrP properties in a variety of assays, but we still do not clearly understand the structural correlates of PrP toxicity. Additional insight can be extracted from comparative structural studies, followed by molecular dynamics simulations of selected mutations, and testing in manipulable animal models. Our working hypothesis is that protective amino acids generate more compact and stable structures in a C-terminal subdomain of the PrP globular domain. We will explore this idea in this review and identify subdomains within the globular domain that may hold the key to unravel how conformational stability and disease susceptibility are encoded in PrP.

D159 and S167 are protective residues in the prion protein from dog and horse, two prion-resistant animals

Prion diseases are fatal neurodegenerative diseases caused by misfolding of the prion protein (PrP). These conditions affect humans and animals, including endemic forms in sheep and deer. Bovine, rodents, and many zoo mammals also developed prion diseases during the “mad-cow” epidemic in the 1980's. Interestingly, rabbits, horses, and dogs show unusual resistance to prion diseases, suggesting that specific sequence changes in the corresponding endogenous PrP prevents the accumulation of pathogenic conformations. In vitro misfolding assays and structural studies have identified S174, S167, and D159 as the key residues mediating the stability of rabbit, horse, and dog PrP, respectively. Here, we expressed the WT forms of rabbit, horse, and dog PrP in transgenic Drosophila and found that none of them is toxic. Replacing these key residues with the corresponding amino acids in hamster PrP showed that mutant horse (S167D) and dog (D159N) PrP are highly toxic, whereas mutant rabbit (S174 N) PrP is not. These results confirm the impact of S167 and D159 in local and long-range structural features in the globular domain of PrP that increase its stability, while suggesting the role of additional residues in the stability of rabbit PrP. Identifying these protective amino acids and the structural features that stabilize PrP can contribute to advance the field towards the development of therapies that halt or reverse the devastating effects of prion diseases.

In Vitro Approach To Identify Key Amino Acids in Low Susceptibility of Rabbit Prion Protein to Misfolding.

Prion diseases, or transmissible spongiform encephalopathies (TSEs), are a group of rare progressive neurodegenerative disorders caused by an abnormally folded prion protein (PrPSc). This is capable of transforming the normal cellular prion protein (PrPC) into new infectious PrPSc Interspecies prion transmissibility studies performed by experimental challenge and the outbreak of bovine spongiform encephalopathy that occurred in the late 1980s and 1990s showed that while some species (sheep, mice, and cats) are readily susceptible to TSEs, others are apparently resistant (rabbits, dogs, and horses) to the same agent. To study the mechanisms of low susceptibility to TSEs of certain species, the mouse-rabbit transmission barrier was used as a model. To identify which specific amino acid residues determine high or low susceptibility to PrPSc propagation, protein misfolding cyclic amplification (PMCA), which mimics PrPC-to-PrPSc conversion with accelerated kinetics, was used. This allowed amino acid substitutions in rabbit PrP and accurate analysis of misfolding propensities. Wild-type rabbit recombinant PrP could not be misfolded into a protease-resistant self-propagating isoform in vitro despite seeding with at least 12 different infectious prions from diverse origins. Therefore, rabbit recombinant PrP mutants were designed to contain every single amino acid substitution that distinguishes rabbit recombinant PrP from mouse recombinant PrP. Key amino acid residue substitutions were identified that make rabbit recombinant PrP susceptible to misfolding, and using these, protease-resistant misfolded recombinant rabbit PrP was generated. Additional studies characterized the mechanisms by which these critical amino acid residue substitutions increased the misfolding susceptibility of rabbit PrP.IMPORTANCE Prion disorders are invariably fatal, untreatable diseases typically associated with long incubation periods and characteristic spongiform changes associated with neuronal loss in the brain. Development of any treatment or preventative measure is dependent upon a detailed understanding of the pathogenesis of these diseases, and understanding the mechanism by which certain species appear to be resistant to TSEs is critical. Rabbits are highly resistant to naturally acquired TSEs, and even under experimental conditions, induction of clinical disease is not easy. Using recombinant rabbit PrP as a model, this study describes critical molecular determinants that confer this high resistance to transmissible spongiform encephalopathies.

Prion protein dynamics before aggregation

Prion diseases, like Alzheimer's disease and Parkinson disease, are rapidly progressive neurodegenerative disorders caused by misfolding followed by aggregation and accumulation of protein deposits in neuronal cells. Here we measure intramolecular polypeptide backbone reconfiguration as a way to understand the molecular basis of prion aggregation. Our hypothesis is that when reconfiguration is either much faster or much slower than bimolecular diffusion, biomolecular association is not stable, but as the reconfiguration rate becomes similar to the rate of biomolecular diffusion, the association is more stable and subsequent aggregation is faster. Using the technique of Trp-Cys contact quenching, we investigate the effects of various conditions on reconfiguration dynamics of the Syrian hamster and rabbit prion proteins. This protein exhibits behavior in all three reconfiguration regimes. We conclude that the hamster prion is prone to aggregation at pH 4.4 because its reconfiguration rate is slow enough to expose hydrophobic residues on the same time scale that bimolecular association occurs, whereas the rabbit sequence avoids aggregation by reconfiguring 10 times faster than the hamster sequence.

Unique Properties of the Rabbit Prion Protein Oligomer.

Prion diseases, also known as transmissible spongiform encephalopathies (TSEs), are a group of fatal neurodegenerative disorders infecting both humans and animals. Recent works have demonstrated that the soluble prion protein oligomer (PrPO), the intermediate of the conformational transformation from the host-derived cellular form (PrPC) to the disease-associated Scrapie form (PrPSc), exerts the major neurotoxicity in vitro and in vivo. Rabbits show strong resistance to TSEs, the underlying mechanism is unclear to date. It is expected that the relative TSEs-resistance of rabbits is closely associated with the unique properties of rabbit prion protein oligomer which remain to be addressed in detail. In the present work, we prepared rabbit prion protein oligomer (recRaPrPO) and human prion protein oligomer (recHuPrPO) under varied conditions, analyzed the effects of pH, NaCl concentration and incubation temperature on the oligomerization, and compared the properties of recRaPrPO and recHuPrPO. We found that several factors facilitated the formation of prion protein oligomers, including low pH, high NaCl concentration, high incubation temperature and low conformational stability of monomeric prion protein. RecRaPrPO was formed more slowly than recHuPrPO at physiological-like conditions (< 57°C, < 150 mM NaCl). Furthermore, recRaPrPO possessed higher susceptibility to proteinase K and lower cytotoxicity in vitro than recHuPrPO. These unique properties of recRaPrPO might substantially contribute to the TSEs-resistance of rabbits. Our work sheds light on the oligomerization of prion proteins and is of benefit to mechanistic understanding of TSEs-resistance of rabbits.

Investigation on salt bridge interactions of mammalian prion proteins by molecular dynamics simulation / Memeli prion proteinlerinin moleküler dinamik simulasyon ile tuz köprüleri etkileşimlerinin araştırılması

Abstract Objective: Salt bridge interaction is one of the most important electrostatic interactions to stabilize the secondary and tertiary structures of protein. To obtain more insight into the molecular basis of prion proteins, the salt bridge networks in two animal prion proteins are studied in this work. Methods: Molecular dynamics (MD) and Flow MD (FMD) simulations are employed to investigate the salt bridges interactions of rabbit prion protein (rPrPc), Syrian hamster prion protein (syPrPc) and the variants of the two prion proteins. Results: The dynamic behaviors of salt bridges are characterized, and the relation between salt bridge interactions and local structures are also discussed. The type of salt bridges in the two prion proteins is divided into the helixloop, intra-helix and inter-helix salt bridges. It is found that the helix-loop salt bridges is more important for the stability of prion proteins than the other two kinds of slat bridge. Conclusion: The Asp201-Arg155 (rS1), Asp177-Arg163 (rS3) and Asp178-Arg164 (syS1) are the important salt bridges to stabilize the structures of rPrPc and syPrPc, respectively. The structural stability is partly depended on the number of helix-loop salt bridge.

Protein misfolding cyclic amplification induces the conversion of recombinant prion protein to PrP oligomers causing neuronal apoptosis.

The formation of neurotoxic prion protein (PrP) oligomers is thought to be a key step in the development of prion diseases. Recently, it was determined that the sonication and shaking of recombinant PrP can convert PrP monomers into β-state oligomers. Herein, we demonstrate that β-state oligomeric PrP can be generated through protein misfolding cyclic amplification from recombinant full-length hamster, human, rabbit, and mutated rabbit PrP, and that these oligomers can be used for subsequent research into the mechanisms of PrP-induced neurotoxicity. We have characterized protein misfolding cyclic amplification-induced monomer-to-oligomer conversion of PrP from three species using western blotting, circular dichroism, size-exclusion chromatography, and resistance to proteinase K (PK) digestion. We have further shown that all of the resulting β-oligomers are toxic to primary mouse cortical neurons independent of the presence of PrP(C) in the neurons, whereas the corresponding monomeric PrP were not toxic. In addition, we found that this toxicity is the result of oligomer-induced apoptosis via regulation of Bcl-2, Bax, and caspase-3 in both wild-type and PrP(-/-) cortical neurons. It is our hope that these results may contribute to our understanding of prion transformation within the brain. We found that β-state oligomeric PrPs can be generated through protein misfolding cyclic amplification (PMCA) from recombinant full-length hamster, human, rabbit, and mutated rabbit PrP. β-oligomers are toxic to primary mouse cortical neurons independent of the presence of PrP(C) in the neurons, while the corresponding monomeric PrPs were not toxic. This toxicity is the result of oligomers-induced apoptosis via regulation of Bcl-2, Bax, and caspase-3. These results may contribute to our understanding of prion transformation within the brain.

Aggregation Propensity of Prion is Kinetically Controlled by Intramolecular Diffusion of Protein Chain

Prion disease are rapidly progressive neurodegenerative disorders like Alzheimer's disease and Parkinson disease that are attributed to misfolding followed by ordered aggregation and accumulation of protein deposits in neuronal cells. According to the “protein only” hypothesis, the central event in prion pathogenesis is the conformational conversion of the cellular α-helical rich isoform, PrPC, into misfolded β-sheet rich isoform, PrPSc. Here we present a novel way to understand the molecular basis of Prion misfolding and aggregation, by investigating the early stage intramolecular backbone reconfiguration of monomer. Our hypothesis is that when reconfiguration is much faster or much slower than bimolecular diffusion, biomolecular association is not stable, but as the reconfiguration rate becomes similar to the rate of biomolecular diffusion, the association is more stable and subsequent aggregation is faster. We are investigating the effects of reducing agent, pH and denaturant on reconfiguration dynamics of the Hamster and Rabbit Prion protein. This work not only provides a better understanding of early stage conformational dynamics, but also provides a way forward for a novel therapeutic strategy of drug development which can bind monomers and prevent aggregation at the earliest step.

Molecular dynamics studies on the NMR structures of rabbit prion protein wild type and mutants: surface electrostatic charge distributions

Prion diseases are invariably fatal and highly infectious neurodegenerative diseases that affect a wide variety of mammalian species such as sheep and goats, cattle, deer and elk, and humans. But for rabbits, studies have shown that they have a low susceptibility to be infected by prion diseases. This paper does molecular dynamics (MD) studies of rabbit NMR structures (of the wild type and its two mutants of two surface residues), in order to understand the specific mechanism of rabbit prion proteins (RaPrPC). Protein surface electrostatic charge distributions are specially focused to analyze the MD trajectories. This paper can conclude that surface electrostatic charge distributions indeed contribute to the structural stability of wild-type RaPrPC; this may be useful for the medicinal treatment of prion diseases.

COMPARING FOLDING MECHANISMS OF DIFFERENT PRION PROTEINS BY Gō MODEL

Prions are associated with neurodegenerative diseases induced by transmissible spongiform encephalopathies. The infectious scrapie form is referred to as PrP Sc , which has conformational change from normal prion with predominant α-helical conformation to the abnormal PrP Sc that is rich in β-sheet content. Neurodegenerative diseases have been found from both human and bovine sources, but there are no reports about infected by transmissible spongiform encephalopathies from rabbit, canine and horse sources. Here we used coarse-grained Gō model to compare the difference among human, bovine, rabbit, canine, and horse normal (cellular) prion proteins. The denatured state of normal prion has relation with the conversion from normal to abnormal prion protein, so we used all-atom Gō model to investigate the folding pathway and energy landscape for human prion protein. Through using coarse-grained Gō model, the cooperativity of the five prion proteins was characterized in terms of calorimetric criterion, sigmoidal transition, and free-energy profile. The rabbit and horse prion proteins have higher folding free-energy barrier and cooperativity, and canine prion protein has slightly higher folding free-energy barrier comparing with human and bovine prion proteins. The results from all-atom Gō model confirmed the validity of C α-Gō model. The correlations of our results with previous experimental and theoretical researches were discussed.

Decipher the mechanisms of rabbit's low susceptibility to prion infection

Rabbits have low susceptibility to prion infection. Studies on prion protein (PrP) from animal species of different susceptibility to prion diseases identified key amino acid residues, specific motif, and special features in rabbit prion protein (RaPrP(C)) that contribute to the stability of rabbit PrP(C) and low susceptibility to prion infection. However, there is no evidence showing that rabbits are completely resistant to prion diseases. It has been reported that the rabbit prion could be generated in vitro through protein misfolding cyclic amplification and proved to be infectious and transmissible. Here, we reviewed studies on rabbit-specific PrP structures and features in relation to rabbit's low susceptibility to prion infection.

Molecular dynamics studies on the NMR and X-ray structures of rabbit prion proteins

Prion diseases, traditionally referred to as transmissible spongiform encephalopathies (TSEs), are invariably fatal and highly infectious neurodegenerative diseases that affect a wide variety of mammalian species, manifesting as scrapie in sheep and goats, bovine spongiform encephalopathy (BSE or mad-cow disease) in cattle, chronic wasting disease in deer and elk, and Creutzfeldt–Jakob diseases, Gerstmann–Sträussler–Scheinker syndrome, fatal familial insomnia, and kulu in humans, etc. These neurodegenerative diseases are caused by the conversion from a soluble normal cellular prion protein (PrPC) into insoluble abnormally folded infectious prions (PrPSc), and the conversion of PrPC to PrPSc is believed to involve conformational change from a predominantly α-helical protein to one rich in β-sheet structure. Such a conformational change may be amenable to study by molecular dynamics (MD) techniques. For rabbits, classical studies show that they have a low susceptibility to be infected by PrPSc, but recently it was reported that rabbit prions can be generated through saPMCA (serial automated Protein Misfolding Cyclic Amplification) in vitro and the rabbit prion is infectious and transmissible. In this paper, we first do a detailed survey on the research advances of rabbit prion protein (RaPrP) and then we perform MD simulations on the NMR and X-ray molecular structures of rabbit prion protein wild-type and mutants. The survey shows to us that rabbits were not challenged directly in vivo with other known prion strains and the saPMCA result did not pass the test of the known BSE strain of cattle. Thus, we might still look rabbits as a prion resistant species. MD results indicate that the three α-helices of the wild-type are stable under the neutral pH environment (but under low pH environment the three α-helices have been unfolded into β-sheets), and the three α-helices of the mutants (I214V and S173N) are unfolded into rich β-sheet structures under the same pH environment. In addition, we found an interesting result that the salt bridges such as ASP201–ARG155, ASP177–ARG163 contribute greatly to the structural stability of RaPrP.

3D local structure around copper site of rabbit prion-related protein: Quantitative determination by XANES spectroscopy combined with multiple-scattering calculations

Prion-related protein (PrP), a cell-surface copper-binding glycoprotein, is considered to be responsible for a number of transmissible spongiform encephalopathies (TSEs). The structural conversion of PrP from the normal cellular isoform (PrPC) to the post-translationally modified form (PrPSc) is thought to be relevant to Cu2+ binding to histidine residues. Rabbits are one of the few mammalian species that appear to be resistant to TSEs, because of the structural characteristics of the rabbit prion protein (RaPrPC) itself. Here we determined the three-dimensional local structure around the C-terminal high-affinity copper-binding sites using X-ray absorption near-edge structure combined with ab initio calculations in the framework of the multiple-scattering (MS) theory. Result shows that two amino acid resides, Gln97 and Met108, and two histidine residues, His95 and His110, are involved in binding this copper(II) ion. It might help us understand the roles of copper in prion conformation conversions, and the molecular mechanisms of prion-involved diseases.

The Contrasting Effect of Macromolecular Crowding on Amyloid Fibril Formation

BackgroundAmyloid fibrils associated with neurodegenerative diseases can be considered biologically relevant failures of cellular quality control mechanisms. It is known that in vivo human Tau protein, human prion protein, and human copper, zinc superoxide dismutase (SOD1) have the tendency to form fibril deposits in a variety of tissues and they are associated with different neurodegenerative diseases, while rabbit prion protein and hen egg white lysozyme do not readily form fibrils and are unlikely to cause neurodegenerative diseases. In this study, we have investigated the contrasting effect of macromolecular crowding on fibril formation of different proteins.Methodology/Principal FindingsAs revealed by assays based on thioflavin T binding and turbidity, human Tau fragments, when phosphorylated by glycogen synthase kinase-3β, do not form filaments in the absence of a crowding agent but do form fibrils in the presence of a crowding agent, and the presence of a strong crowding agent dramatically promotes amyloid fibril formation of human prion protein and its two pathogenic mutants E196K and D178N. Such an enhancing effect of macromolecular crowding on fibril formation is also observed for a pathological human SOD1 mutant A4V. On the other hand, rabbit prion protein and hen lysozyme do not form amyloid fibrils when a crowding agent at 300 g/l is used but do form fibrils in the absence of a crowding agent. Furthermore, aggregation of these two proteins is remarkably inhibited by Ficoll 70 and dextran 70 at 200 g/l.Conclusions/SignificanceWe suggest that proteins associated with neurodegenerative diseases are more likely to form amyloid fibrils under crowded conditions than in dilute solutions. By contrast, some of the proteins that are not neurodegenerative disease-associated are unlikely to misfold in crowded physiological environments. A possible explanation for the contrasting effect of macromolecular crowding on these two sets of proteins (amyloidogenic proteins and non-amyloidogenic proteins) has been proposed.

The Structural Stability of Wild-type Horse Prion Protein

Prion diseases (e.g. Creutzfeldt-Jakob disease (CJD), variant CJD (vCJD), Gerstmann-Sträussler-Scheinker syndrome (GSS), Fatal Familial Insomnia (FFI) and Kuru in humans, scrapie in sheep, bovine spongiform encephalopathy (BSE or ‘mad-cow’ disease) and chronic wasting disease (CWD) in cattles) are invariably fatal and highly infectious neurodegenerative diseases affecting humans and animals. However, by now there have not been some effective therapeutic approaches or medications to treat all these prion diseases. Rabbits, dogs, and horses are the only mammalian species reported to be resistant to infection from prion diseases isolated from other species. Recently, the β2-α2 loop has been reported to contribute to their protein structural stabilities. The author has found that rabbit prion protein has a strong salt bridge ASP177-ARG163 (like a taut bow string) keeping this loop linked. This paper confirms that this salt bridge also contributes to the structural stability of horse prion protein. Thus, the region of β2-α2 loop might be a potential drug target region. Besides this very important salt bridge, other four important salt bridges GLU196-ARG156-HIS187, ARG156-ASP202 and GLU211-HIS177 are also found to greatly contribute to the structural stability of horse prion protein. Rich databases of salt bridges, hydrogen bonds and hydrophobic contacts for horse prion protein can be found in this paper.

Molecular Dynamics Studies on the Structural Stability of Wild-type Dog Prion Protein

Prion diseases such as Creutzfeldt-Jakob disease, variant Creutzfeldt-Jakob diseases, Gerstmann-Sträussler-Scheinker syndrome, Fatal Familial Insomnia, Kuru in humans, scrapie in sheep, bovine spongiform encephalopathy (or ‘mad-cow’ disease) and chronic wasting disease in cattle are invariably fatal and highly infectious neurodegenerative diseases affecting humans and animals. However, by now there have not been some effective therapeutic approaches to treat all these prion diseases. In 2008, canine mammals including dogs (canis familials) were the first time academically reported to be resistant to prion diseases (Vaccine 26: 2601-2614 (2008)). Thus, it is very worth studying the molecular structures of dog prion protein to obtain insights into the immunity of dogs to prion diseases. This paper studies the molecular structural dynamics of wild-type dog prion protein. The comparison analyses with rabbit prion protein show that the dog prion protein has stable molecular structures whether under neutral or low pH environments. We also find that the salt bridges such as D177-R163 contribute to the structural stability of wild-type rabbit prion protein under neutral pH environment.

Progresses on prion proteins

Transmissible spongiform encephalopathies are a family of fatal neurodegenerative disorders.The pathogen has been identified as the abnormal isoform of the host-derived prion protein.The molecular mechanism of the conformational conversion of the prion protein is unclear so far.However,much valuable information closely related to the conformational change has been obtained by means of physical,chemical and biological techniques. Recent progresses on three-dimensional structures,dynamics,unfolding and intermolecular interactions of prion proteins are reviewed herein,together with novel results about the solution structure and dynamics of the rabbit prion protein achieved from our laboratory.

Sequence-dependent Prion Protein Misfolding and Neurotoxicity

Prion diseases are neurodegenerative disorders caused by misfolding of the normal prion protein (PrP) into a pathogenic "scrapie" conformation. To better understand the cellular and molecular mechanisms that govern the conformational changes (conversion) of PrP, we compared the dynamics of PrP from mammals susceptible (hamster and mouse) and resistant (rabbit) to prion diseases in transgenic flies. We recently showed that hamster PrP induces spongiform degeneration and accumulates into highly aggregated, scrapie-like conformers in transgenic flies. We show now that rabbit PrP does not induce spongiform degeneration and does not convert into scrapie-like conformers. Surprisingly, mouse PrP induces weak neurodegeneration and accumulates small amounts of scrapie-like conformers. Thus, the expression of three highly conserved mammalian prion proteins in transgenic flies uncovered prominent differences in their conformational dynamics. How these properties are encoded in the amino acid sequence remains to be elucidated.

Prion disease susceptibility is affected by β-structure folding propensity and local side-chain interactions in PrP

Prion diseases occur when the normally α-helical prion protein (PrP) converts to a pathological β-structured state with prion infectivity (PrP(Sc)). Exposure to PrP(Sc) from other mammals can catalyze this conversion. Evidence from experimental and accidental transmission of prions suggests that mammals vary in their prion disease susceptibility: Hamsters and mice show relatively high susceptibility, whereas rabbits, horses, and dogs show low susceptibility. Using a novel approach to quantify conformational states of PrP by circular dichroism (CD), we find that prion susceptibility tracks with the intrinsic propensity of mammalian PrP to convert from the native, α-helical state to a cytotoxic β-structured state, which exists in a monomer-octamer equilibrium. It has been controversial whether β-structured monomers exist at acidic pH; sedimentation equilibrium and dual-wavelength CD evidence is presented for an equilibrium between a β-structured monomer and octamer in some acidic pH conditions. Our X-ray crystallographic structure of rabbit PrP has identified a key helix-capping motif implicated in the low prion disease susceptibility of rabbits. Removal of this capping motif increases the β-structure folding propensity of rabbit PrP to match that of PrP from mouse, a species more susceptible to prion disease.