Recombinant Escherichia coli cell for D-p-hydroxyphenylglycine production from D-N-carbamoyl-p-hydroxyphenylglycine

Recombinant Escherichia coli cell containing D-amidohydrolase was employed to convert D-N-carbamoyl-p-hydroxyphenylglycine (D-CpHPG) to D-p-hydroxyphe nylglycine (D-pHPG). Biotransformations under pH 7 and 40 degrees C allowed to complete conversion of D-CpHPG into D-pHPG. Under the same reaction DH, the D-amidohydrolase activity of the cell in the phosphate buffer was high er than that in the Tris buffer. The activity decreased with the increase o f phosphate buffer. concentration, instead of using buffer, the reaction pH maintained constant at 7 by titrating with 1 N HCl resulted in a higher D- pHPG production rate. Flocculating the cell suspension with chitosan and cr oss-linked by glutaraldehyde mode the cell recovery for repeated use much e asier. Both the cross-linking and (PMSF; a protease inhibitor) treatments c ould increase the cell reusability and storage stability. However, the cros s-linking decreased the D-amidohydrolase activity of the cell to about 50%. The D-amidohydrolase activities of fret: and cross-linked cell were: inhib ited at substrate concentration higher than 150 mM and 100 mM, respectively . The conversion of 150 mM D-CpHPG to D-pHPG could be completed within 7 h for the free cell at the concentration of 10% (wet weight/volume). (C) 2000 Elsevier Science Inc. All rights reserved.