Semisynthetic prion protein (PrP) variants carrying glycan mimics at position 181 and 197 do not form fibrils

Abstract

Semisynthesis and characterization of homogeneously mono- and di-PEGylated full length PrP variants to study the impact of PEGylation (as N-glycan mimics) on protein folding and aggregation.

Prion protein—Semisynthetic prion protein (PrP) variants with posttranslational modifications

“…Thus, lipopeptides can be used as mimics of the GPI anchor's ability to attach PrP to the cell membrane. A similar strategy with regard to using a lipidated peptide as a GPI‐mimicking membrane anchor was pursued by Becker et al, when starting to develop semisynthetic strategies, based on EPL and protein trans ‐splicing (PTS) (Figure ), to access to different posttranslationally modified PrP variants (Figure ) …”

“…This strategy was aimed at introduction sugars into PrP but did not fully succeed. Shortly after that, Araman et al demonstrated a semisynthetic approach to generate PrP variants modified with monodisperse PEG chains as mimics of N ‐glycans that are similar in size and molar mass (Figure C). A new EPL strategy was established to achieve this, in which expressed PrP (aa 23‐177) with a C ‐terminal thioester is used in EPL reactions with PEGylated, synthetic peptides (aa 179‐231).…”

“…Different strategies were utilized by Baskakov, 252 Baldwin, 253 Figure 4), to access to different posttranslationally modified PrP variants ( Figure 5). 228,241,[255][256][257][258][259][260][261] In the EPL 245,246 reaction, a protein thioester, obtained by cleaving a fusion protein consisting of the protein of interest (POI) and an intein, is linked to a chemically synthesized peptide 243 containing an N-terminal cysteine in a reaction similar to NCL. 244 An initial transesterifaction leads to formation of a thioester linking the recombinant and synthetic PrP segments, and a subsequent irreversible S → N acyl shift establishes the amide bond at the ligation site ( Figure 4A).…”

Building Peptide Bonds in Haifa: The Seventh Chemical Protein Synthesis (CPS) Meeting

“…Christian Becker (University of Vienna, Austria) discussed a semisynthetic approach to generate prion protein (PrP) variants modified with monodisperse poly(ethylene glycol) (PEG) units as mimics of N‐glycans. The incorporation of PEG at different glycosylation sites in PrP only induced small changes in the secondary structure, but effectively inhibited the aggregation of PrP in vitro …”

“…The incorporation of PEG at different glycosylation sites in PrP only induceds mall changesi n the secondary structure,b ut effectivelyi nhibited the aggregation of PrP in vitro. [26] Apart from glycoproteins, recent efforts in understanding PTMs such as SUMOylation, ubiquitylation, phosphorylation, and lipidation of proteins were also discussed. Champak Chatterjee (University of Washington, USA) presented his laboratory's efforts towards understanding the mechanistic basis of gene regulation by chromatin SUMOylation.…”