Prion protein self-peptides modulate prion interactions and conversion

<p>Abstract</p> <p>Background</p> <p>Molecular mechanisms underlying prion agent replication, converting host-encoded cellular prion protein (PrP^C) into the scrapie associated isoform (PrP^Sc), are poorly understood. Selective self-interaction between PrP molecules forms a basis underlying the observed differences of the PrP^Cinto PrP^Scconversion process (agent replication). The importance of previously peptide-scanning mapped ovine PrP self-interaction domains on this conversion was investigated by studying the ability of six of these ovine PrP based peptides to modulate two processes; PrP self-interaction and conversion.</p> <p>Results</p> <p>Three peptides (octarepeat, binding domain 2 -and C-terminal) were capable of inhibiting self-interaction of PrP in a solid-phase PrP peptide array. Three peptides (N-terminal, binding domain 2, and amyloidogenic motif) modulated prion conversion when added before or after initiation of the prion protein misfolding cyclic amplification (PMCA) reaction using brain homogenates. The C-terminal peptides (core region and C-terminal) only affected conversion (increased PrP^resformation) when added before mixing PrP^Cand PrP^Sc, whereas the octarepeat peptide only affected conversion when added after this mixing.</p> <p>Conclusion</p> <p>This study identified the putative PrP core binding domain that facilitates the PrP^C-PrP^Scinteraction (not conversion), corroborating evidence that the region of PrP containing this domain is important in the species-barrier and/or scrapie susceptibility. The octarepeats can be involved in PrP^C-PrP^Scstabilization, whereas the N-terminal glycosaminoglycan binding motif and the amyloidogenic motif indirectly affected conversion. Binding domain 2 and the C-terminal domain are directly implicated in PrP^Cself-interaction during the conversion process and may prove to be prime targets in new therapeutic strategy development, potentially retaining PrP^Cfunction. These results emphasize the importance of probable PrP^C-PrP^Cand required PrP^C-PrP^Scinteractions during PrP conversion. All interactions are probably part of the complex process in which polymorphisms and species barriers affect TSE transmission and susceptibility.</p

Prion protein self-peptides

Molecular mechanisms underlying prion agent replication, converting host-encoded cellular prion protein (PrPC) into the scrapie associated isoform (PrPSc), are poorly understood. Selective self-interaction between PrP molecules forms a basis underlying the observed differences of the PrPC into PrPSc conversion process (agent replication). The importance of previously peptide-scanning mapped ovine PrP self-interaction domains on this conversion was investigated by studying the ability of six of these ovine PrP based peptides to modulate two processes, PrP self-interaction and conversion

Prion Protein Self-Peptides

Molecular mechanisms underlying prion agent replication, converting host-encoded cellular prion protein (PrPC) into the scrapie associated isoform (PrPSc), are poorly understood. Selective self-interaction between PrP molecules forms a basis underlying the observed differences of the PrPC into PrPSc conversion process (agent replication). The importance of previously peptide-scanning mapped ovine PrP self-interaction domains on this conversion was investigated by studying the ability of six of these ovine PrP based peptides to modulate two processes, PrP self-interaction and conversion

Computer-Assisted Analysis and Epidemiological Value of Genotyping Methods for Campylobacter jejuni and Campylobacter coli

For epidemiological tracing of the thermotolerant Campylobacter species C. jejuni and C. coli, reliable and highly discriminatory typing techniques are necessary. In this study the genotyping techniques of flagellin typing (flaA typing), pulsed-field gel electrophoresis (PFGE), automated ribotyping, and amplified fragment length polymorphism (AFLP) fingerprinting were compared. The following aspects were compared: computer-assisted analysis, discriminatory power, and use for epidemiological typing of campylobacters. A set of 50 campylobacter poultry isolates from The Netherlands and neighboring countries was analyzed. Computer-assisted analysis made cluster analysis possible and eased the designation of different genotypes. AFLP fingerprinting was the most discriminatory technique, identifying 41 distinct genotypes, while PFGE identified 38 different types, flaA typing discriminated 31 different types, and ribotyping discriminated 26 different types. Furthermore, AFLP analysis was the most suitable method for computer-assisted data analysis. In some cases combining the results of AFLP fingerprinting, PFGE, and flaA typing increased our ability to differentiate strains that appeared genetically related. We conclude that AFLP is a highly discriminatory typing method and well suited for computer-assisted data analysis; however, for optimal typing of campylobacters, a combination of multiple typing methods is needed

Amplified Fragment Length Polymorphism Analysis of Campylobacter jejuni Strains Isolated from Chickens and from Patients with Gastroenteritis or Guillain-Barré or Miller Fisher Syndrome

The high-resolution genotyping method of amplified fragment length polymorphism (AFLP) analysis was used to study the genetic relationships between Campylobacter jejuni strains infecting chickens (n = 54) and those causing gastroenteritis in humans (n = 53). In addition, C. jejuni strains associated with the development of Guillain-Barré syndrome (GBS) (n = 14) and Miller Fisher syndrome (MFS) (n = 4), two related acute paralytic syndromes in human, were included. Strains were isolated between 1989 and 1998 in The Netherlands. The AFLP banding patterns were analyzed with correlation-based and band-based similarity coefficients and UPGMA (unweighted pair group method using average linkages) cluster analysis. All C. jejuni strains showed highly heterogeneous fingerprints, and no fingerprints exclusive for chicken strains or for human strains were obtained. All strains were separated in two distinct genetic groups. In group A the percentage of human strains was significantly higher and may be an indication that genotypes of this group are more frequently associated with human diseases. We conclude that C. jejuni from chickens cannot be distinguished from human strains and that GBS or MFS related strains do not belong to a distinct genetic group

sH5³ vaccination of mice.

<p>Groups of 10 BALB/c mice were immunized i.m. with 2 µg sH5³ either once (day 21) or twice with a 3-week interval (day 0 and day 21). As a challenge control, one group of mice was mock-treated (PBS) twice (day 0 and day 21). Three weeks after the vaccination, mice were infected with ∼10 LD₅₀ of A/Viet Nam/1194/04 and monitored daily for clinical signs and body weight during the next 14 days. (A) Kaplan-Meier survival curve indicating percentage mortality on each day for each group. (B) Median clinical scores per group. (C) Mean body weights per group expressed as percentage of starting body weight, plotted as a function of time. Error bars represent the standard deviation. (D–E) Blood samples were collected at the day of challenge. sH5³ antibody levels as determined by ELISA (D). HI titers against sH5³ (E). Bars represent geometric means.</p