In biochemistry, PMSF (phenylmethanesulfonylfluoride or phenylmethylsulfonyl fluoride) is a serine protease inhibitor commonly used in the preparation of cell lysates. PMSF does not inhibit all serine proteases. It is rapidly degraded in water and stock solutions are usually made up in anhydrous ethanol, isopropanol, corn oil, or DMSO. Proteolytic inhibition occurs when a concentration between 0.1 – 1 mM PMSF is used. The half-life is short in aqueous solutions (110 min at pH=7 and 35 min at pH=8).
PMSF binds specifically to the active site serine residue in a serine protease, but does not bind to any other serine residues in the protein. Since PMSF binds covalently to the enzyme at the active serine residue, the complex can be viewed by X-ray crystallography; it can therefore be used as a chemical label to identify an essential active site SER in an enzyme.
- Enzyme (active) Ser-O-H + F-SO2CH2C6H5 -> EnzymeSer-O-SO2CH2C6H5 + HF
Serine protease + PMSF -> Irreversible enzyme-PMS complex + HF
PMSF is a cytotoxic chemical that should only be handled inside a fume hood; the LD50 for this compound is less than 500 mg/kg.
Preparation of PMSF (10 mM), 10 ml
- Weigh out 17.4 mg PMSF
- Add isopropanol to 10ml and dissolve.
- No need to filter sterilize.
- Aliquot and store at –20°C.
PMSF is unstable in aqueous solutions with a half-life of approximately 30 minutes. Add solution immediately before use.
PMSF: Sigma (P-7626)