When prions are transmitted between species, there can be a delay in pathogenesis due to a phenomenon referred to as the transmission barrier. Some species also show very low susceptibility to prion disease. In this study I hypothesized that the susceptibility of species to prion disease is proportional to the tendency of their endogenous prion protein, PrP, to adopt the β-state, an oligomeric form of misfolded recombinant PrP that is rich in β sheet. Using a novel method of two-wavelength CD analysis, it could be shown that recombinant PrP from prion-susceptible species have a higher propensity to refold to the β-state than resistant species. The crystal structure of rabbit PrPC 121-230 revealed a helix-cap motif at the N-terminus of helix-2 that contributes to the reduced β-state propensity of rabbit PrP. Single amino acid changes in the sequence of PrP can lead to a transmission barrier and/or resistance in species. Mutating single residues in rabbit PrPC to those found in corresponding positions in hamster PrPC, ablated the helix-cap observed in the wild-type and caused an increase in the β-state propensity of rabbit PrP. Conversely, a decrease in β-state propensity was observed when rabbit mutations were introduced into PrP of hamster, a susceptible species. A dimeric association is hypothesized to be involved in the function of PrP and/or the conversion mechanism to infectious prion. In the structures of the wild-type and mutant rabbit PrPCs a dimeric arrangement was observed in the asymmetric unit of the crystals. Using 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC), a dimer of rabbit and hamster PrPC was crosslinked in solution. The dimer crosslink was specific and dependent on the tertiary structure of PrPC. Crosslinking of the β-state octamer with EDC showed that similar contacts may be present in this oligomeric form. Together these data provide strong evidence that species susceptibility is linked to β-state propensity.

When prions are transmitted between species, there can be a delay in pathogenesis due to a phenomenon referred to as the transmission barrier. Some species also show very low susceptibility to prion disease. In this study I hypothesized that the susceptibility of species to prion disease is proportional to the tendency of their endogenous prion protein, PrP, to adopt the β-state, an oligomeric form of misfolded recombinant PrP that is rich in β sheet. Using a novel method of two-wavelength CD analysis, it could be shown that recombinant PrP from prion-susceptible species have a higher propensity to refold to the β-state than resistant species. The crystal structure of rabbit PrPC 121-230 revealed a helix-cap motif at the N-terminus of helix-2 that contributes to the reduced β-state propensity of rabbit PrP. Single amino acid changes in the sequence of PrP can lead to a transmission barrier and/or resistance in species. Mutating single residues in rabbit PrPC to those found in corresponding positions in hamster PrPC, ablated the helix-cap observed in the wild-type and caused an increase in the β-state propensity of rabbit PrP. Conversely, a decrease in β-state propensity was observed when rabbit mutations were introduced into PrP of hamster, a susceptible species. A dimeric association is hypothesized to be involved in the function of PrP and/or the conversion mechanism to infectious prion. In the structures of the wild-type and mutant rabbit PrPCs a dimeric arrangement was observed in the asymmetric unit of the crystals. Using 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC), a dimer of rabbit and hamster PrPC was crosslinked in solution. The dimer crosslink was specific and dependent on the tertiary structure of PrPC. Crosslinking of the β-state octamer with EDC showed that similar contacts may be present in this oligomeric form. Together these data provide strong evidence that species susceptibility is linked to β-state propensity.

Transmissible spongiform encephalopathies (TSEs), or prion diseases, are a group of invariably fatal neurodegenerative diseases of both humans and animals, thought to be caused by the abnormally folded prion protein PrPSc. Prion disease research continues to be faced by a number of difficult challenges. First, the unequivocal diagnosis of most prion diseases currently requires the post-mortem collection of central nervous system tissue, either for histological examination or Western blot analysis; second, a viable treatment for clinical stage disease has not yet been identified; third, the exact details of disease pathogenesis have not been elucidated; and fourth, the normal function of PrPC is not definitively known. The primary objective of the studies presented here was to diagnose prion disease in live animals, using Magnetic Resonance Imaging (MRI). Increases in T2 relaxation time and apparent diffusion coefficient (ADC) were observed very early following the infection of Syrian golden hamsters with the 263K strain of scrapie. These changes were evident well before the appearance of either clinical symptoms or the typical histological changes characteristic of prion disease, suggesting that they are the result of the progressive accumulation of fluid, and that this may constitute a novel early marker of prion disease pathogenesis. Following the establishment of this model system, a secondary objective was composed: to test the viability of a potential treatment (pentosan polysulphate) using a number of different treatment regimens. It was determined that pentosan polysulphate (PPS) was ineffective as a treatment unless it was administered intra-cerebrally very early in infection, although it was shown to slow the appearance of the histological hallmarks of prion disease. In response to the results of these studies, a potential model was proposed, relating PrP, aquaporin-4 (AQP4) regulation, and oedema. Although speculative, this model may have implications for both normal PrPC function and disease pathogenesis.

Multi-component reaction (MCR) provides an easy and rapid access to libraries of organic compounds with diverse substitution patterns. This project aims to employ MCR for synthesis of both combinatorial (CL) and dynamic combinatorial (DCL) libraries for lead discovery.

Transmissible spongiform encephalopathies (TSEs), or prion diseases, are a group of invariably fatal neurodegenerative diseases of both humans and animals, thought to be caused by the abnormally folded prion protein PrPSc. Prion disease research continues to be faced by a number of difficult challenges. First, the unequivocal diagnosis of most prion diseases currently requires the post-mortem collection of central nervous system tissue, either for histological examination or Western blot analysis; second, a viable treatment for clinical stage disease has not yet been identified; third, the exact details of disease pathogenesis have not been elucidated; and fourth, the normal function of PrPC is not definitively known. The primary objective of the studies presented here was to diagnose prion disease in live animals, using Magnetic Resonance Imaging (MRI). Increases in T2 relaxation time and apparent diffusion coefficient (ADC) were observed very early following the infection of Syrian golden hamsters with the 263K strain of scrapie. These changes were evident well before the appearance of either clinical symptoms or the typical histological changes characteristic of prion disease, suggesting that they are the result of the progressive accumulation of fluid, and that this may constitute a novel early marker of prion disease pathogenesis. Following the establishment of this model system, a secondary objective was composed: to test the viability of a potential treatment (pentosan polysulphate) using a number of different treatment regimens. It was determined that pentosan polysulphate (PPS) was ineffective as a treatment unless it was administered intra-cerebrally very early in infection, although it was shown to slow the appearance of the histological hallmarks of prion disease. In response to the results of these studies, a potential model was proposed, relating PrP, aquaporin-4 (AQP4) regulation, and oedema. Although speculative, this model may have implications for both normal PrPC function and disease pathogenesis.

Zsfassung in engl. Sprache. - Würzburg, Univ., Diss., 2008

Thesis (Ph. D.)--Washington State University, December 2008. / Title from PDF title page (viewed on Oct. 22, 2009). "College of Veterinary Medicine." Includes bibliographical references.

Thesis (Ph. D.)--University of Chicago, Dept. of Molecular Genetics and Cell Biology. / Includes bibliographical references. Also available on the Internet.

Dissertation (Ph.D.) --University of Texas Graduate School of Biomedical Sciences at San Antonio, 2008. / Vita. Includes bibliographical references.

Würzburg, Univ., Diss., 2009.