Bioconversion of D-HPG Using Immobilized Genetic Engineered Strain HC01

Abstract:

The cells of engineered strain HC01 were immobilized in the form of Ca2+-alginate beads. The conditions for immobilization were investigated. The optimal gel concentration and cell concentration were found to be 2.5% and 0.029 g/mL in the presence of 3% CaCl2. The thermo-stability of immobilized cells was 5°C higher than that of free cells in the same condition. Divalent metal ions, such as Mn2+, Mg2+, Cu2+, Co2+ and Ni2+ did not affect significantly the enzymatic activity of D-hydantoinase (HYD) and N-carbamoylase (CAB) in immobilized cells. By contrast, Mn2+ and Ni2+ could independently enhance the activity of CAB up to 210% and 270% in free cells. The present data showed that the optimal reaction condition of immobilized cells was at pH 7.5 and 40°C as same as that of free cells. The immobilized cells were applied to produce D-p-Hydroxyphenylglycine (D-HPG) directly from the substrate DL-Hydroxyphenyl Hydantoin (DL-HPH) in a batch reactor. Conversion of about 97% was reached after 36-h reaction when the substrate concentration was 30 g/L with the initial pH of 9.0 under the protection of nitrogen gas. The overall yield of D-HPG was 85% with the optical purity of 99.7% after purification.

Key words:D-Amino acid, Engineering strain, Immobilization, Calcium alginate