3.3 Solubilizing the inclusion body in Urea
According to theSDS-PAGE analysis, inclusion bodies of FPOX were only
soluble in urea 8 M with a slight solubilization in the presence of 4 M
urea by traditional method while was not soluble in any of used
concentrations of DMSO solvent (Fig.3). In freeze-thaw method, the
inclusion bodies were not soluble in urea 0.5 M while were soluble in
urea 1 and 2 M with a concentration of 1mg/mL and 1.4 mg/mL,
respectively, while it was not soluble with these concentrations of urea
in traditional method. However, the major part of inclusion bodies of
FPOX was soluble in urea 8 M by freeze-thaw method. The dialysis method
was used to refold soluble inclusion bodies; which gradually permitted
the alteration from denaturing to native buffer conditions. However,
considerable precipitation was observed after dilution to the refolding
buffer without urea. Supernatant containing refolded FPOX was collected
after centrifugation, and used for the activity assay.