Proteinase K (PROK)

Proteinase K (EC 3.4.21.64) is a highly active, broad-spectrum serine protease originally isolated from the fungus Engyodontium album [UniProt: P06873]. It belongs to the subtilisin-like family and is characterized by its ability to cleave peptide bonds at the carboxylic sides of aliphatic, aromatic, or hydrophobic amino acids [PubChem: SID 472051441]. The enzyme is notably stable and active across a wide range of pH values and temperatures, and it retains activity in the presence of detergents such as SDS, which makes it an essential tool in molecular biology for the digestion of proteins during nucleic acid purification [PubMed: 2385944]. While Proteinase K is not a therapeutic target for human disease, it is widely used in research and diagnostics, particularly for the detection of the protease-resistant infectious prion protein (PrPSc) in transmissible spongiform encephalopathies [PubMed: 10859301]. In laboratory settings, its activity can be inhibited by serine protease inhibitors such as phenylmethylsulfonyl fluoride (PMSF) or diisopropyl fluorophosphate (DFP), which act by covalently modifying the catalytic serine residue [Wikipedia: Proteinase K].

Other names

Endopeptidase KTritirachium alkaline proteinaseEC 3.4.21.64

Inhibitors such as phenylmethylsulfonyl fluoride (PMSF) and diisopropyl fluorophosphate (DFP) target Proteinase K by covalently binding to the active site serine residue (Ser224), thereby irreversibly inactivating the enzyme's catalytic triad and preventing proteolysis.