Title of article :
Renaturation, purification and characterization of streptokinase expressed as inclusion body in recombinant E. coli
Author/Authors :
Cherish Babu، نويسنده , , P.V. and Srinivas، نويسنده , , V.K. and Krishna Mohan، نويسنده , , V. and Krishna، نويسنده , , Ella، نويسنده ,
Issue Information :
روزنامه با شماره پیاپی سال 2008
Pages :
9
From page :
218
To page :
226
Abstract :
Recombinant protein purification is facilitated using high expression systems which produce larger quantities of streptokinase protein as inclusion bodies. As the accumulation of active streptokinase is toxic to the host cells, we have optimized the conditions to achieve large amounts of streptokinase in the form of inclusion bodies. The solubility and yield of pure protein are highly dependent on various combinations of chemical additives, ionic and non-ionic detergents and salts, with solubilizing agents followed by refolding of denatured protein into active form. As the extraction of the purified streptokinase from inclusion bodies requires denaturation and a subsequent refolding step, careful balancing steps were needed to develop under different controlled conditions. Here the purified fragments of refolded proteins were screened to select the conditions that yield the active streptokinase having native conformation. The maximum specific activity of the purified streptokinase was achieved by these methods. The refolded recombinant streptokinase was analyzed by RP-HPLC showing a purity of 99%. Size exclusion chromatography profile shows that there are minimal aggregates in the active streptokinase protein and the percentage of renaturation is around 99%.
Keywords :
Physiochemical characterization , Recombinant streptokinase (SK) , Escherichia coli fermentation , Solubilization , Inclusion body isolation , Protein Purification
Journal title :
Journal of Chromatography B
Serial Year :
2008
Journal title :
Journal of Chromatography B
Record number :
1465551
Link To Document :
بازگشت