Degradation and detection of TSE infectivity
View Patent ↗A transmissible spongiform encephalopathy (TSE) agent is inactivated by exposing the TSE agent to a thermostable proteolytic enzyme at elevated temperature and at acid or alkaline pH. Following this step, or separately, presence of TSE infectivity is detected by detection of dimers of prion protein.
1. A method for inactivating a transmissible spongiform encephalopathy (TSE) agent, located on or within infected apparatus or materials, or in clinical waste, said method comprising exposing the TSE agent to a thermostable proteolytic enzyme at an alkaline pH of from 8 to 13,
wherein the exposing of the TSE agent to the thermostable proteolytic enzyme is carried out at a temperature that is 45° C. to less than 100° C., and
the TSE agent is a prion.
2. The method of claim 1 , wherein the pH is from 10 to 12.
3. The method of claim 1 , wherein the TSE agent results in a disease is selected from the group consisting of Creutzfeld-Jacob disease; variant Creutzfeld-Jacob disease; Kuru; fatal familial insomnia; Gerstmann-Straussler-Scheinker syndrome; bovine spongiform encephalopathy; scrapie; feline spongiform encephalopathy; chronic wasting disease; and transmissible mink encephalopathy.
4. The method of claim 1 , wherein the thermostable proteolytic enzyme is obtained from a thermophilic organism.
5. The method of claim 4 , wherein the thermophilic organism is selected from the group consisting of Thermotoga maritima; Thermotoga neopolitana; Thermotoga thermarum; Fervidobacterium islandicum; Fervidobacterium nodosum; Fervidobacterium pennivorans; Thermosipho africanus; Aeropyrum pernix; Thermus flavus; pyrococcus spp.; Sulfolobus solfataricus; Desulfurococcus; Bacillus thermoproteolyticus; Bacillus stearo - thermophilus; Bacillus sp. 11231 ; Bacillus sp. 11276 ; Bacillus sp. 11652 ; Bacillus sp. 12031 ; Thermus aquaticus; Thermus caldophilus; Thermus sp. 16132 ; Thermus sp. 15673; and Thermus sp. Rt41A.
6. The method of claim 4 wherein the thermophilic organism is selected from the group consisting of archaea; hyperthermophilic bacteria and thermophilic bacteria.
7. The method of claim 1 , wherein the thermostable proteolytic enzyme is selected from the group consisting of MC-A, MC-3 and MC-4.
8. The method of claim 1 , wherein the pH is from 9 to 12.
9. The method of claim 1 , wherein the pH is about 12.
10. The method of claim 1 , wherein the exposing of the TSE agent to the thermostable proteolytic enzyme is carried out at a temperature that is about 60° C. to about 80° C.
11. The method of claim 1 , wherein the TSE agent is a prion or a prion dimer.
12. The method of claim 1 , further comprising exposing the TSE agent to the thermostable proteolytic enzyme and a detergent.
13. A method of claim 1 , further comprising exposing the TSE agent to the thermostable proteolytic enzyme and an enzyme selected from the group consisting of proteases, lipases, peptidases, glycosylases, and nucleases.
14. A method of claim 1 , further comprising exposing the TSE agent to the thermostable proteolytic enzyme and sonication.
15. The method of claim 1 , wherein the temperature is 50° C. to less than 100° C.
16. The method of claim 15 , wherein the temperature is between 55° C. and 85° C.
17. The method of claim 1 , wherein the thermostable proteolytic enzyme is a serine protease.
18. The method of claim 1 , wherein the thermostable proteolytic enzyme is a subtilisin.
19. The method of claim 18 , wherein the thermostable proteolytic enzyme is a subtilisin derived from Bacillus bacteria.
20. The method of claim 19 wherein the thermostable proteolytic enzyme is a subtilisin derived from Bacillus amyloliquefaciens, Bacillus lentus, Bacillus licheniformis, Bacillus subtilis or is subtilisin PB92.