Inhibition of prion amplification by expression of dominant inhibitory mutants--a systematic insertion mutagenesis study.
Until now it is still not clear which structural elements of the prion protein (PrP) are involved in its conversion process. Characterisation of these essential regions would help to understand the conversion process itself and might help to develop specific therapeutic approaches to inhibit PrP(res) formation by dominant inhibitory mutations. To address this important question 33 evenly spaced insertion mutants were generated spanning the entire sequence of the murine 3F4-tagged PrP. The mutants were expressed by retroviral transduction in three different scrapie infected cell lines (ScN2a; SMB[RC040]; SMB[22F]). The convertibility was affected not only by introducing the insertion in the putatively refolded region (aa100-170), but also in the C-terminus of PrP (up to aa214). Moreover, dominant inhibitory effects on conversion were observed for PrP-mutants at four distinguished regions (aa100-112; aa130-154; aa166-172, aa196-200). Computer based structural analysis revealed that these segments were organized in two structurally clearly separated regions supporting the idea that they could function as protein-protein interaction sites which are necessary during seed formation.
Geissen
Markus
M
Institute for Novel and Emerging Infectious Diseases at the Friedrich-Loeffler-Institut, Greifswald-Insel Riems, Germany. martin.groschup@fli.bund.de
Mella
Harriet
H
Saalmüller
Armin
A
Eiden
Martin
M
Proft
Juliane
J
Pfaff
Eberhard
E
Schätzl
Hermann M
HM
Groschup
Martin H
MH
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D005786:Gene Expression Regulation; D006031:Glycosylation; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D018620:PrPSc Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D054730:Protein Interaction Domains and Motifs; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D021381:Protein Transport
D018620:PrPSc Proteins; D020033:Protein Isoforms