3.3 Optimization of reaction conditions for whole-cell biotransformation
Outstanding thermostability of SI has always been pursued in successful industrial manufacturing bioprocess for isomaltulose, as even slight enhancement can improve long-term activity under optimum conditions, increase the ability to remain high activity in repeat batches of whole-cell biotransformation. To develop an economically feasible whole-cell biocatalysis process, the influence of temperature, pH, cell density as variables were explored. As shown in Figure 3A, the optimum pH of whole-cell activity was 6.0, which was higher than that of pure enzyme expressed from C. glutamicum . Wu’s report also showed the same shift in the optimal pH, which may be attributed by the differences of micro-environment.[23] Whereas no obvious changes were detected in the optimum temperature between free enzyme and whole-cell activity, all the maximum catalytic activity was at 30℃ (Figure 3B). Moreover, cell dosage also plays an important in the actual production, and reaction time is related to the cell density during the biotransformation process. Figure 3C shows that the concentration of isomaltulose was highest in the first 1.5 h when the cell density was OD600=30, while the yield of isomaltulose at OD600=25 was significantly higher than OD600=30 after 1.5 h. Therefore, OD600=25 was selected as the optimal cell dosage for the synthesis of isomaltulose.