title: Production of α-cyclodextrin glycosyltransferase in Bacillus megaterium MS941 by systematic codon usage optimization.
abstract: α-Cyclodextrin glycosyltransferase is a key enzyme in the cyclodextrin industry. The Gram-positive bacterium Bacillus megaterium was chosen for production of recombinant α-CGTase for safety concerns. Successful production of heterologous α-CGTase was achieved by adapting the original α-cgt gene to the codon usage of B. megaterium by systematic codon optimization. This balanced the tRNA pool and reduced ribosomal traffic jams. Protein expression and secretion was ensured by using the strong inducible promoter P(xyl) and the signal peptide SP(LipA). The impact of culture medium composition and induction strategies on α-CGTase production was systematically analyzed. Production and secretion at 32 °C for 24 h using modified culture medium was optimal for α-CGTase yield. Batch- and simple fed-batch fermentation was applied to achieve a high yield of 48.9 U·mL(-1), which was the highest activity reported for a Bacillus species, making this production system a reasonable alternative to Escherichia coli.
authors.affiliation: School of Biotechnology and Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi, Jiangsu, China. zhoujw1982@jiangnan.edu.cn
authors.forename: Jingwen
authors.initials: J
authors.lastname: Zhou
authors.forename: Hua
authors.initials: H
authors.lastname: Liu
authors.forename: Guocheng
authors.initials: G
authors.lastname: Du
authors.forename: Jianghua
authors.initials: J
authors.lastname: Li
authors.forename: Jian
authors.initials: J
authors.lastname: Chen
mesh_terms: D001410:Bacillus megaterium / Q000201:enzymology* / Q000235:genetics*; D003062:Codon / Q000235:genetics; D003470:Culture Media; D005285:Fermentation; D015870:Gene Expression; D005964:Glucosyltransferases / Q000096:biosynthesis* / Q000235:genetics; D005990:Glycerol / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000235:genetics; D021382:Protein Sorting Signals; D011994:Recombinant Proteins / Q000096:biosynthesis*
chemical_list: D003062:Codon; D003470:Culture Media; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D005964:Glucosyltransferases; C019963:cyclomaltodextrin glucanotransferase; D005990:Glycerol