Journal
BIORESOURCE TECHNOLOGY
Volume 172, Issue -, Pages 174-179Publisher
ELSEVIER SCI LTD
DOI: 10.1016/j.biortech.2014.09.035
Keywords
Pullulanase; Escherichia coli; Extracellular secretion; Glycine; Glycine feeding strategy
Funding
- National Natural Science Foundation of China [31271813, 31100048]
- Natural Science Foundation of Jiangsu Province [BK20140142, BK20140132]
- 111 Project [111-2-06]
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Process optimization strategies were developed to improve extracellular production of recombinant Bacillus deramificans pullulanase in Escherichia coli. Cell growth and pullulanase production in shake-flask cultures were investigated as a function of the concentration of added glycine, and the type and concentration of inducer. From the results of these experiments, a fed-batch fermentation strategy for high-cell-density cultivation was applied in a 3-L fermentor. The gradual addition of lactose was utilized for the induction of protein expression. The optimal lactose feeding rate and induction point were 0.4 g L-1 h(-1) and a dry cell weight (DCW) of 15 g L-1, respectively. Furthermore, a glycine feeding strategy was formulated to promote the secretion of recombinant protein. The optimal total and extracellular pullulanase activity were 2523.5 and 1567.9 U mL(-1), respectively, which represent 1.2 and 22.6-fold increases compared with those observed under unoptimized conditions. (C) 2014 Elsevier Ltd. All rights reserved.
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