Immobilization of a multi enzyme system for L amino acids production
BACKGROUND Four enzymes were immobilized for the production of optically pure natural and non natural L amino acids via the 'Double Racemase Hydantoinase Process' Immobilization constitutes an empirical process, and for each enzyme we tested 11 carriers with different functional groups; epoxide, EC EP, EC HFA, IB 150 and IB 350, carboxylic acid IB C435, quaternary ammonium IB A161, IB A171 and IB A369, aromatic group IB S861, and hydroxyl group IB S60S and IB S60P. RESULTS Each protein showed preference for binding on one or several supports: D,L hydantoinase/IB 350, hydantoin racemase/EC EP, L carbamoylase/EC EP and carbamoyl racemase/IB A161. The process was optimized for each enzyme by modifying temperature, pH and ionic strength. For the enzymatic cascade, it was demonstrated that it was essential to use supports having homogeneous characteristics. The product of the first reaction is the substrate of the next one, and so on. Free mass diffusion from one enzyme to the other is crucial to avoid retention on the support and to maintain the protein substrate interaction constant. CONCLUSION It was proved that support size, weight and hydrophobicity must be homogeneous to avoid the formation of separate layers of the matrix selected. For this reason, the support IB 350 was used for the four enzymes, even though it may not be the most efficient one for some of them. (c) 2015 Society of Chemical Industry