Hydantoinases/dihydropyrimidinases are important biotechnological enzymes involved in the production of α- and β-amino acids. Their isolation from new sources with different substrate specificities, improved activity, enantioselectivity, or higher stability continues to be of great industrial interest. Here, we provide a detailed description of how to produce high quantities of the recombinant hydantoinase/dihydropyrimidinase enzyme from Sinorhizobium meliloti CECT4114 (SmeDhp). Several techniques are combined to obtain this goal, from cloning to activity measurement by HPLC.