Subtilisin is a non-specific protease (a protein-digesting enzyme) initially obtained from Bacillus subtilis.

Subtilisins belong to subtilases, a group of serine proteases that - like all serine proteases - initiate the nucleophilic attack on the peptide (amide) bond through a serine residue at the active site. Subtilisins typically have molecular weights of about 20,000 to 45,000 dalton. They can be obtained from certain types of soil bacteria, for example, Bacillus amyloliquefaciens from which they are secreted in large amounts.