Characterization of a proteolytic enzyme derived from a Bacillus strain that effectively degrades prion protein

Abstract

Aims: The purpose of this paper was to screen candidate bacterial strains for the production of proteases suitable for application to the degradation of pathogenic forms of prion protein (PrPSc). This paper describes the biochemical characteristics and proteolytic activity of the isolated protease. Methods and Results: After screening more than 200 bacterial proteases for keratinolytic activity, we identified a Bacillus stain that produced a protease exhibiting high‐degradation activity against a scrapie PrPSc. Sequence analysis indicated that this serine‐protease belonged to the Subtilisin family and had optimum pH and temperature ranges of 9–10 and 60–70°C. Western blotting analysis revealed that the protease was also capable of decomposing bovine spongiform encephalopathy‐infected brain homogenate. In addition, the protease was demonstrated to degrade dried PrPSc that had become firmly attached to a plastic surface considerably more effectively than proteinase K or PWD‐1, a previously reported keratinase. Conclusions: These results indicate that the isolated protease exhibited higher activity for PrPSc degradation compared with other proteases examined. Significance and Impact of the Study: This protease could be used under moderate conditions for the decontamination of precision instruments that are susceptible to PrPSc contamination.

A Review: Biodegradation and Applications of Keratin Degrading Microorganisms and Keratinolytic Enzymes, Focusing on Thermophiles and Thermostable Serine Proteases

“…The temperature and pH optima in the presence of CaCl 2 were about 105°C and pH 6.5 (Šnajder et al, 2012). Purified keratinase from the strain MSK103 was also capable of degrading and decontaminating prion infected brain homogenate at 50°C without detergents or heat pretreatment (Yoshioka et al, 2007). The MSK103 enzyme had strong activity between 60-70°C and optimal pH was 9-10.…”

“…BA7, Actinomadura keratinilytica Cpt29 and Nocardiopsis sp. TOA-1 (Williams et al, 1990;Lin et al, 1992;Letourneau et al, 1998;Wang and Shih, 1999;Korkmaz et al, 2003;Gousterova et al, 2005;Mitsuiki et al, 2006;Yoshioka et al, 2007;Habbeche et al, 2014).…”

Potential application spectrum of microbial proteases for clean and green industrial production

“…Proteases may help degrading wrongly folded proteins. Yoshioka et al (2007) identified a protease-producing Bacillus strain that was capable of degrading scrapie PrP Sc . The protease (MSK 103) was also effective against dried PrP Sc .…”

Prions in the environment

“…Bacterial enzymes which effectively degrade prions have been identified, but they are most effective at high pH (10-12) and high temperature (50-60°C). 59,104 Microbiological consortia taken from the rumen and colon of cattle could degrade PrP Sc to undetectable levels within 20 hours under anaerobic conditions at 37°C, although infectivity remained. 60,105 Using an enzyme treatment, it may be possible to lower or eliminate the infectivity at identified or presumed CWD and scrapie 'hot spots' in captive and wild settings.…”