Improved Fmoc‐based solid‐phase synthesis of homologous peptide fragments of human and mouse prion proteins

Abstract

The synthesis of difficult peptide sequences has been a challenge since the very beginning of SPPS. The self-assembly of the growing peptide chains has been proposed as one of the causes of this synthetic problem. However, there is an increasing need to obtain peptides and proteins that are prone to aggregate. These peptides and proteins are generally associated with diseases known as amyloidoses. We present an efficient SPPS of two homologous peptide fragments of HuPrP (106-126) and MoPrP105-125 based on the use of the PEGA resin combined with proper coupling approaches. These peptide fragments were also studied by CD and TEM to determine their ability to aggregate. On the basis of these results, we support PEG-based resins as an efficient synthetic tool to prepare peptide sequences prone to aggregate on-resin.

Neurotoxicity of Prion Peptides Mimicking the Central Domain of the Cellular Prion Protein

“…These results suggest that cytotoxicity might be exerted by the unstable spherical species of the peptide detected by TEM (Figure 2C), which disappear over time after resuspension (Figures 2G and 2K). As expected from previous results (e.g., [19]), PrP 106–126 showed significant toxicity when aggregated for 24 h after dissolution (Figure 3B), supporting the delayed appearance of toxic aggregated species in this peptide with time [53]. Lastly, we performed a time course with different concentrations of the CD peptide for different periods of incubation.…”

“…Several in vitro studies have shown the high dynamics and reversibility in the equilibrium between monomer and protofibril formation of other amyloid proteins (e.g., Aβ [52]), and the coexistence of different species due to polymorphic fibril assembly pathways [66]. Regarding PrP 106–126 , the equilibrium between monomers and soluble oligomers, with an enrichment in secondary structures, is independent of concentration [34], but the fibrillar forms increase with time in a progressive manner [53] in parallel with the toxic intermediate structures. Taken together, the different described effects of PrP 106–126 could be attributed to the particular experimental conditions in each study.…”

Selenomethionine Incorporation into Amyloid Sequences Regulates Fibrillogenesis and Toxicity

“…For its part, [SeM 109,112,129,134 ]HuPrP(106–140) (all-M), its all-Val counterpart and the four site-specifically SeM-substituted analogs were efficiently assembled by microwave-assisted solid phase synthesis on ChemMatrix®, a resin proven successful in preventing aggregation during the synthesis of large, complex peptides [43]. Peptides were successfully purified from the crude material and their identity and homogeneity confirmed by mass spectrometry [44]. Full details on the synthesis, purification and analytical documentation of all peptides are given in the Supporting Information S1 file.…”

Functional recruitment of dynamin requires multimeric interactions for efficient endocytosis

“…Couplings of Fmoc-protected amino acids (0.2 M) were carried out in the presence of HBTU (0.25 M) and DIPEA (1M) using standard coupling cycles (20 W, 70°C, 300s, except for Fmoc-Arg(Pbf)-OH (0 W, RT, 1500s followed by 20 W, 70°C, 300s). For the synthesis of D44, we evaluated a priori difficult couplings according to (Grillo-Bosch et al, 2011). We thus introduced a systematic double coupling on Y1, A2, L3, G5 (N-terminus amino acids) V37, P38, I40 and T41 (bulky amino acids).…”