| Bioactivity | Proteinase K (Protease K) is a nonspecific serine protease that is useful for general digestion of proteins. Proteinase K is active in the presence of SDS or urea and over a wide range of pH (4-12), salt concentrations, and temperatures. Proteinase K can be use for promoting methods of viral nucleic acid extraction, and detection[1][2][3][4]. | ||||||||||||
| Invitro | Proteinase K enzyme can be used to break amid bond between MTX and BSA and also amidic bonds in BSA structure[2].Guidelines (Following is our recommended protocol. This protocol only provides a guideline, and should be modified according to your specific needs). Proteinase K with superior protein-degrading property, can be used to detect the SARS-COV2 viral component directly from inoculated viral transport medium (VTM) without RNA extraction[3]. Protocol of rRTPCR detection of SARS-CoV2[3]:1.Separate VTM of each SARS-COV2 samples into three EP tubes (45 μL per tube);2.Add 2.5 µL (20 mg/mL concentrated) proteinase K (PK), making a final concentration of PK ~1 mg/ml VTM;3.Incubate at 25 ℃ for 5 min;4.Incubate at 60 ℃ for 15 min;5.Incubate at 98 ℃ for 10 min, and store samples at 4 ℃;6.Use 5 µL of extracted RNA as template to execute rRTPCR assay;7.Exert rRTPCR detection by BIORAD CFX96™ Real-Time Thermocycler, with a positive standard of threshold cycle (Ct) values less than 40.Proteinase K improves the homogenization effects and positive rates of sputum samples, and increases nucleic acid concentration extracted from it[4].Protocol of nucleic acid extraction of influenza A virus (IAV)[4]:1.Separate specimen into three EP tubes (500 μL per tube);2.Add 500 μL 0.4 mg/mL Proteinase K buffer, vortex for 20 seconds;3.Incubate at 55 ℃ for 25 min, vortex 5 seconds every 5 min;4.Extract RNA by using the Automated Nucleic Acid Extraction System through a paramagnetic beads method (Zhijiang Biotechnology Co., Ltd., Shanghai, China) with 200 μL samples;5.Evaluate A260/A280 ratio of pure RNA products, with a standard ranged from 1.8 to 2.0. | ||||||||||||
| Name | Proteinase K | ||||||||||||
| CAS | 39450-01-6 | ||||||||||||
| Appearance | Solid | ||||||||||||
| Transport | Room temperature in continental US; may vary elsewhere. | ||||||||||||
| Storage |
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| Reference | [1]. W Ebeling, et al. Proteinase K From Tritirachium Album Limber. Eur J Biochem . 1974 Aug 15;47(1):91-7. [2]. Nosrati H, et al. Multifunctional nanoparticles from albumin for stimuli-responsive efficient dual drug delivery. Bioorg Chem. 2019 Jul;88:102959. [3]. Shukla A, et al. Vitality of Proteinase K in rRTPCR Detection of SARS-CoV2 Bypassing RNA Extraction. Front Cell Infect Microbiol. 2021 Nov 3;11:717068. [4]. Yu F, et al. Comparative Evaluation of Three Preprocessing Methods for Extraction and Detection of Influenza A Virus Nucleic Acids from Sputum. Front Med (Lausanne). 2018 Mar 2;5:56. |