Stadnyk V. The biochemical characteristics of prion-protein in normal conditions and pathology

The thesis is devoted to investigation of the biochemical particularities of cellular and pathological prions and their isoforms in normal conditions and in the animal prion infections. The biochemical features of the cellular prion and its isoforms have been established in different structural and functional parts of cattle and laboratory rats brain. It was showed that the highest level of this protein expression is observed in medulla oblongata, olfactory tract and olfactory bulb. In other parts of cattle cerebrum, namely in epiphysis, brain hemispheres, inferior colliculus and cerebellum cellular prion protein is also expressing, but its level is lower approximately on 30-60% in comparison with medulla oblongata and parts of olfactory tract. Consequently, cellular prion is expressing not only in medulla oblongata but also in other parts of cerebrum which means that these parts of brain can also take part in the pathogenesis of prion diseases. In brain tissue of cattle were detected for the first time the additional isoforms of cellular prion which have intermediate position between di- and deglycosilated prion isoforms. It was shown, that in a case of atypical form of BSE, pathological prion accumulates in olive nucleus of brainstem in diffuse manner. Typical form of BSE was characterized by granular accumulation of prion aggregates in intercellular space of dorsal nucleus of brainstem. It was decided to make a search of preparations which are able to decrease a level of cellular prion with out side effects, and in the same time depriving pathological prion of substrate for replication. Were chosen two compounds - heparin and pentosan polysulfate, as they are able to interact with cellular prion in vitro. Speaking about the influence of these compounds on cellular prion in vivo we must mark that there is no such reference data. The results of our investigations show that injections of heparin and pentosan polysulphate cause decrease of cellular prion level in forebrain, brainstem and spinal cord of tested animals. It was set that type of PrPc isoforms expression after PPS treatment changes in forebrain, were take place decrese of deglycosylated form of cellular prion as a result of decrease of level of diglycosilated PrPc and also in the brainstem, were PPs causes expression of monoglycosylated form of cellular prion, which could not be detected in normal condirions. In the same time we found out that injection of PPS causes considerable increase of copper and zinc level in tested tissues when activity of Cu/Zn-SOD does not change. Heparin in concentrations 1 and 50 mg/kg/day does not cause changes in the maintenance of copper and zinc, and in activity of Cu/Zn-SOD. Although injections with heparin in dose 300 mg/kg/day leads to the decrease of Cu/Zn-SOD activity but has no influence on levels of copper and zinc. Molecular mechanism of interaction between cellular recombinant prion and pentosan polysulphate has been investigated. It was detected, that this interaction is specific, has high-affinity and is provided by non-covalent binding sulphate groups of pentosan polysulphate with Arg148 and Arg151 amino acid residues in prion polypeptide chain. These results can explain the inhibiting effect of pentosan polysulphate on prion protein expression in vivo. Key words: prion protein, isoforms of prion, heparin, pentosan polysulphate, bovine spongiform encephalopathy.