|
KMID : 0545120080180101648
|
|
Journal of Microbiology and Biotechnology
2008 Volume.18 No. 10 p.1648 ~ p.1654
|
|
|
Increased Yield of High-Purity and Active Tetrameric Recombinant Human EC-SOD by Solid Phase Refolding
|
|
Ryu Kang
Kim Young-Hoon
Kim Young-Hwa
Lee Joon-Seok
Jeon Byeong-Wook
Kim Tae-Yoon
|
|
|
Abstract
|
|
|
|
Superoxide dismutase (SOD) removes damaging reactive oxygen species from the cellular environment by catalyzing the dismutation of two superoxide radicals to hydrogen peroxide and oxygen. Extracellular superoxide dismutase (EC-SOD) is a tetramer and is present in the extracellular space and to a lesser extent in the extracellular fluids. Increasing therapeutic applications for recombinant human extracellular superoxide dismutase (rEC-SOD) has broadened interest in optimizing methods for its purification, with a native conformation of tetramer. We describe a solid phase refolding procedure that combines immobilized metal affinity chromatography (IMAC) and gel filtration chromatography in the purification of rEC-SOD from Escherichia coli. The purified rEC-SOD tetramer from the Ni2+-column chromatography is refolded in Tris buffer. This method yields greater than 90% of the tetramer form. Greater than 99% purity is achieved with further purification over a Superose 12PC 3.2/30 column to obtain the tetramer and specific activities as determined via DCFHDA assay. The improved yield of rEC-SOD in a simple chromatographic purification procedure promises to enhance the development and therapeutic application of this biologically potent molecule.
|
|
|
KEYWORD
|
|
|
Extracellular superoxide dismutase, solid phase refolding, chromatographic purification procedure
|
|
|
FullTexts / Linksout information
|
|
|
|
|
|
Listed journal information
|
|
  
|