Cloning, purification, crystallization and preliminary structural studies of penicillin V acylase from Bacillus subtilis

Penicillin acylase proteins are amidohydrolase enzymes that cleave penicillins at the amide bond connecting the side chain to their β‐lactam nucleus. An unannotated protein from Bacillus subtilis has been expressed in Escherichia coli, purified and confirmed to possess penicillin V acylase activity. The protein was crystallized using the hanging‐drop vapour‐diffusion method from a solution containing 4 M sodium formate in 100 mM Tris–HCl buffer pH 8.2. Diffraction data were collected under cryogenic conditions to a spacing of 2.5 Å. The crystals belonged to the orthorhombic space group C2221, with unit‐cell parameters a = 111.0, b = 308.0, c = 56.0 Å. The estimated Matthews coefficient was 3.23 Å3 Da−1, corresponding to 62% solvent content. The structure has been solved using molecular‐replacement methods with B. sphaericus penicillin V acylase (PDB code 2pva) as the search model.