|
KMID : 0545119990090060811
|
|
Journal of Microbiology and Biotechnology
1999 Volume.9 No. 6 p.811 ~ p.819
|
|
|
Characterization of the β-Cyclodextrin Glucanotransferase Gene of Bacillus firmus var.alkalophilus and Its Expression in E.coli
|
|
Park, Tae Hyung
Shin, Hyun Dong/Lee, Yong Hyun
|
|
|
Abstract
|
|
|
|
The β-CGTase gene of alkalophilic Bacillus firmus var. alkalophilus was cloned into E. coli using pZErO^TM-2 as a vector. The cloned gene encoded a total of 710 amino acid residues consisting of 674 amino acids of the matured protein and 36 amino acids of the signal peptide, including 20 amino acids from the lacZ gene in the vector. Although the cloned β-CGTase gene did not contain the promoter and start codons, it was expressed by the lac promoter and lacZ start codon in the pZErO^TM-2 vector. A comparison was made with the amino acid sequence and ten other CGTases from Bacillus sp. Also, ten highly conserved regions, which are important amino acid residues in catalysis of CGTase, were identified. The lac promoter used for expression of the β-CGTase gene was induced constitutively in recombinant E. coli even without IPTG possibly because of a lack of the lacl gene in both host and vector, repressing the lacZ gene in the lac operon. Its expression was catabolically repressed by glucose, however, its repression was reduced by soluble starch, mainly because of the extremely high increase of the cAMP level. β-CGTase can be overproduced in the recombinant E. coli by maintaining intracellular cAMP levels mostly through the intermittent feeding of glucose during cultivation.
|
|
|
KEYWORD
|
|
|
|
|
|
FullTexts / Linksout information
|
|
|
|
|
|
Listed journal information
|
|
  
|
|