|
KMID : 0545119980080050509
|
|
Journal of Microbiology and Biotechnology
1998 Volume.8 No. 5 p.509 ~ p.516
|
|
|
Development of a Plasmid Vector for Overproduction of β-Galactosidase in Escherichia coli by Using Genetic Components of groEx from Symbiotic Bacteria in Amoeba proteus
|
|
Lee, Jung Eun
Ahn, Eun Young/Ahn, Tae In
|
|
|
Abstract
|
|
|
|
A plasmid vector, pXGPRMATG-lac-Tgx, was developed for overproduction of β-galactosidase in Escherichia coli using the genetic components of groEx, a heat-shock gene cloned from symbiotic X-bacteria in Amoeba proteus. The vector is composed of intragenic promoters P3 and P4 of groEx, the structural gene of lac operon, transcription terminator signals of lac and groEx, and ColE1 and amp´ of pBluescript SKⅡ. The optimized host, E. coli DH5α, transformed with the vector constitutively produced 117,310-171,961 Miller units of β-galactosidase per mg protein in crude extract. The amount of enzyme in crude extract was 53% of total water-soluble proteins. About 43% of the enzyme could be purified to a specific activity of 322,249 Miller units/㎎ protein after two-fold purification, using two cycles of precipitation with ammonium sulfate and one step of gel filtration. Thus, the expression system developed in this study presents a lowcost and simple method for purifying overproduced β-galactosidase in E. coli.
|
|
|
KEYWORD
|
|
|
|
|
|
FullTexts / Linksout information
|
|
|
|
|
|
Listed journal information
|
|
  
|
|