title: Secretion of cyclodextrin glucanotransferase in E. coli using Bacillus subtilis lipase signal peptide and optimization of culture medium.
abstract: The cyclodextrin glycosyltransferase (CGTase) of Paenibacillus pabuli US132 was fused to the secretive lipase signal peptide of B. subtilis. This leads to an efficient secretion of the recombinant enzyme into the culture medium of E. coli as an active and soluble form contrasting with the native construction leading to a periplasmic production. In order to enhance the yield of CGTase production, an experimental design methodology was applied for the optimization of the culture composition. Hence, the media components were submitted to preliminary screening using a Plakett-Burman design. The concentrations of the major operating ones were then optimized to enhance the secretion of CGTase using response surface methodology. The findings revealed that concentrations of 0.5% potato starch, 3% yeast extract, 3% tryptone, 1.5% casein hydrolysate, 0.5% NaCl, 0.2% KH2PO4, and 0.02% MgSO4 were the optimal conditions for CGTase production. The experimental value (9.43 U/mL) obtained for CGTase activity was very close to the predicted value (9.27 U/mL).
authors.affiliation: Laboratory of Microorganisms and Biomolecules, Centre of Biotechnology of Sfax, Université de Sfax, BP 1177 Sfax, Tunisia. ayadidr@yahoo.fr
authors.forename: Dorra
authors.initials: D
authors.lastname: Ayadi-Zouari
authors.forename: Radhouane
authors.initials: R
authors.lastname: Kammoun
authors.forename: Sonia
authors.initials: S
authors.lastname: Jemli
authors.forename: Hichem
authors.initials: H
authors.lastname: Chouayekh
authors.forename: Samir
authors.initials: S
authors.lastname: Bejar
mesh_terms: D001412:Bacillus subtilis / Q000502:physiology*; D003001:Cloning, Molecular; D003470:Culture Media; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D005964:Glucosyltransferases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008049:Lipase / Q000235:genetics*; D010957:Plasmids; D016133:Polymerase Chain Reaction; D015202:Protein Engineering; D021382:Protein Sorting Signals / Q000235:genetics*; D011994:Recombinant Proteins / Q000378:metabolism*
chemical_list: D003470:Culture Media; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D005964:Glucosyltransferases; C019963:cyclomaltodextrin glucanotransferase; D008049:Lipase