G Liang1, G Du, J Chen. 1. School of Biotechnology, Jiangnan University, Wuxi, China.
Abstract
AIMS: To investigate the effects of pH stress coupled with cysteine addition on glutathione (GSH) production in the treatment of high cell density culture of Candida utilis. METHODS AND RESULTS: We have previously observed that most Candida utilis cells remained viable after being subjected to pH at 1.2 for 3 h and that some intracellular GSH leaked into the medium. A cysteine addition strategy was applied in fed-batch production of GSH. A single cysteine addition resulted in higher GSH yield than two separate additions without pH stress. An increase in intracellular GSH content triggered inhibition of gamma-glutamylcysteine synthetase (gamma-GCS). A strategy that combines cysteine addition with low-pH stress was developed to relieve the inhibition of gamma-GCS. CONCLUSION: Without pH stress, single shot and double shot cysteine addition yielded a total GSH of 1423 and 1325 mg l(-1). In comparison, a low-pH stress counterpart resulted in a total GSH of 1542 and 1730 mg l(-1), respectively. With low-pH stress, we observed GSH secretion into the medium at 673 and 558 mg l(-1) and an increase in the gamma-GCS activity by 1.2- and 1.5-fold, respectively. The specific GSH production yield increased from 1.76% to 1.91% (w/w) for single shot, and 1.64% to 2.14% for double shots. SIGNIFICANCE AND IMPACT OF THE STUDY: Low-pH shift was applied to alleviate the feedback inhibition of intracellular GSH on gamma-GCS activity by secreting GSH into the medium. This strategy is coupled with cysteine addition to enhance GSH production in Candida utilis.
AIMS: To investigate the effects of pH stress coupled with cysteine addition on glutathione (GSH) production in the treatment of high cell density culture of Candida utilis. METHODS AND RESULTS: We have previously observed that most Candida utilis cells remained viable after being subjected to pH at 1.2 for 3 h and that some intracellular GSH leaked into the medium. A cysteine addition strategy was applied in fed-batch production of GSH. A single cysteine addition resulted in higher GSH yield than two separate additions without pH stress. An increase in intracellular GSH content triggered inhibition of gamma-glutamylcysteine synthetase (gamma-GCS). A strategy that combines cysteine addition with low-pH stress was developed to relieve the inhibition of gamma-GCS. CONCLUSION: Without pH stress, single shot and double shot cysteine addition yielded a total GSH of 1423 and 1325 mg l(-1). In comparison, a low-pH stress counterpart resulted in a total GSH of 1542 and 1730 mg l(-1), respectively. With low-pH stress, we observed GSH secretion into the medium at 673 and 558 mg l(-1) and an increase in the gamma-GCS activity by 1.2- and 1.5-fold, respectively. The specific GSH production yield increased from 1.76% to 1.91% (w/w) for single shot, and 1.64% to 2.14% for double shots. SIGNIFICANCE AND IMPACT OF THE STUDY: Low-pH shift was applied to alleviate the feedback inhibition of intracellular GSH on gamma-GCS activity by secreting GSH into the medium. This strategy is coupled with cysteine addition to enhance GSH production in Candida utilis.