title: Delayed supplementation of glycine enhances extracellular secretion of the recombinant alpha-cyclodextrin glycosyltransferase in Escherichia coli.
abstract: The targeting of recombinant proteins for secretion to the culture medium of Escherichia coli presents significant advantages over cytoplasmic or periplasmic expression. However, a major barrier is inadequate secretion across two cell membranes. In the present study, we attempted to circumvent this secretion problem of the recombinant alpha-cyclodextrin glycosyltransferase (alpha-CGTase) from Paenibacillus macerans strain JFB05-01. It was found that glycine could promote extracellular secretion of the recombinant alpha-CGTase for which one potential mechanism might be the increase in membrane permeability. However, further analysis indicated that glycine supplementation resulted in impaired cell growth, which adversely affected overall recombinant protein production. Significantly, delayed supplementation of glycine could control cell growth impairment exerted by glycine. As a result, if the supplementation of 1% glycine was optimally carried out at the middle of the exponential growth phase, the alpha-CGTase activity in the culture medium reached 28.5 U/ml at 44 h of culture, which was 11-fold higher than that of the culture in regular terrific broth medium and 1.2-fold higher than that of the culture supplemented with 1% glycine at the beginning of culture.
authors: State Key Laboratory of Food Science and Technology, Jiangnan University, 1800 Lihu Avenue, Wuxi, People's Republic of China.
authors: Zhaofeng
authors: Z
authors: Li
authors: Zhengbiao
authors: Z
authors: Gu
authors: Miao
authors: M
authors: Wang
authors: Guocheng
authors: G
authors: Du
authors: Jing
authors: J
authors: Wu
authors: Jian
authors: J
authors: Chen
mesh_terms: D019149:Bioreactors; D003470:Culture Media / Q000737:chemistry*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D005964:Glucosyltransferases / Q000378:metabolism*; D005998:Glycine / Q000378:metabolism*; D056507:Paenibacillus / Q000201:enzymology / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism*
chemical_list: D003470:Culture Media; D011994:Recombinant Proteins; D005964:Glucosyltransferases; C019963:cyclomaltodextrin glucanotransferase; D005998:Glycine