Figure 1
From: Recombinant Human Prion Protein Inhibits Prion Propagation in vitro
Inhibition of amplification of human PrPSc by recombinant human PrP (rHuPrP23-231) and mechlorethamine (MCT).
(A) Amplification of human PrPSc from iatrogenic CJD carrying valine (V)/valine polymorphism at residue 129 (129VV) of PrP characteristic of PrPSc type 2 (iCJDVV2, seeds) was carried out by PMCA in the presence of uninfected brain homogenates from humanized transgenic mice expressing PrP-129V (substrates). Lanes 1 and 2: Positive PMCA control without inhibitors; Lanes 3 and 4: PMCA in the presence of 0.2 μM of rHuPrP23-231 with methionine at polymorphic residue 129 (129M); Lanes 5 and 6: PMCA in the presence of 0.2 μM of recombinant human protein disulfide isomerase (rHuPDI); Lanes 1 through 6: PMCA with PrPSc seeds; Lanes 7 and 8: PMCA without PrPSc seeds and inhibitors; Lane 9: The PrP sample without PK-treatment; and Lanes 1 through 8 with PK-treatment. Samples without (−) or with (+) PMCA were treated with 100 μg/ml PK prior to SDS-PAGE and Western blotting with 3F4. Intense PrPSc is detectable in lane 2 (positive control) but not in lane 8 (negative control). However, amplification is virtually undetectable in the presence of rHuPrP23-231 (lane 4) while detectable in the presence of rHuPDI (lane 6). The blot is a representative of five independent experiments. (B) Amplification of human PrPSc from iCJD was carried out in the absence (lanes 3 and 4) or presence (lanes 5 and 6) of MCT. Amplification of PrPSc is inhibited in the presence of MCT (1.5 mM) compared to the sample in the absence of MCT. Lanes 3 through 6 were treated with PK while lanes 1 and 2 were not. The blot is a representative of three independent experiments.
