Study On Conformational Transformation Tendency Of G127V And G131V Mutants Of Human Prion Protein

Prion disease is a fatal neurodegenerative disease whose pathogenesis is closely related to the conversion of cellular prion protein(PrPc)into scrapie prion protein(PrPsc).However,the specific molecular mechanism of prion disease's pathogenesis is still unclear.The key evidence supporting the"protein-only"hypothesis of prion disease lies in the link between hereditary prion disease and the prion protein gene(PRNP).About 10-15%of prion diseases are believed to be associated with genetic mutations.Currently,there are about 40 mutation sites identified in hereditary prion diseases on human PRNP.Among them,the G131V mutant of human prion protein(HuPrP)was identified as a pathogenic mutant of Gerstermann-Straussler-Scheinker syndrome(GSS).On the other hand,fortunately,it is revealed in latest researches that the G127V mutant of HuPrP has strong resistance to Kuru and Creutzefeldt-Jakob diseases(CJD).To provide understanding for the pathogenesis of G131V mutant and the molecular mechanism of disease prevention of G127V mutant,in this thesis,we performed studies on fibrillization and oligomerization propensities for these proteins including wild-type,G127V and G131V mutants of both full-length HuPrP(23-231)and truncated fragment HuPrP(91-231).The experimental results showed that G127V(23-231)has similar fibrillization and oligomerization tendencies to WT(23-231),while the oligomerization tendency of G127V(91?231)is much stronger compared with that of WT(91-231),and the fibrillization tendency is much weaker compared to WT(91-231).The G131V mutation increases the oligomerization tendency of HuPrP and attenuates the fibrillization tendency of HuPrP.In the meantime,we also found that the flexible fragment 23-90 potentiates the oligomerization tendency of WT and the G131V mutant,but attenuates the fibrillization tendency of WT and the G131V mutant In contrast,the fragment 23-90 enhances the fibrillization tendency of the G127V mutant,but decreases the oligomerization tendency of the G127V mutant.The underlying molecular mechaisms deserves further exploring in future.In addition,we also compared the structural and thermodynamic differences of the six HuPrP variants,i.e.WT(23-231),G127V(23-231),G131V(23-231),WT(91-231),G127V(91?231)and G131V(91-231).Previously,our group analyzed the solution structure of the G127V mutant using NMR technology.Compared with the wild type,the 128-131 and 161-164 fragments of G127V(91-231)cannot be assembled into ?-sheets,but forms a Stretch-Strands(SSs)pattern.Compared with the 1H-15N HSQC spectrum of WT(91-231),the shift or broadening of peaks in that of G131V(91-231)is more obvious,indicating that major differences are likely to exist between the spatial structure of WT(91-231)and G131V(91-231).The structural differences of G127V and G131V mutants with the wild type might be the important factors affecting the differences in the conformational transformation tendencies of HuPrP.Furthermore,we measured the thermodynamic stabilities of wild-type and G127V and G131 V mutants by guanidine hydrochloride denaturation and thermal denaturation experiments.We found that G127V and G131V point mutations has no significant effects on structural stabilities of HuPrP,and the effect of the fragment 23-90 on the structural stability of HuPrP is also small.We thus suggest that the thermodynamic stability of the prion protein either is dependent on its conformational transformation tendency with a complicated relationship,or is independent of the tendency.Our results lay a foundation for understanding the conformational transformation of PrP and the pathogenesis of PrP,and provide a theoretical basis for the prevention and cure of prion diseases as well as the design of highly effecient anti-prion drugs.