Abstract
Cystatin C, also known as γ-trace or post-γ-globulin, is a cysteine protease inhibitor from the cystatin superfamily. It is usually used as a marker of the glomerular filtration rate owing to its low molecular weight and constant secretion. The recently available methods for cystatin C preparation have low outputs. Hence, a productive preparation system is urgently required. In this study, a 6 × His-tag coupled with a thrombin cleavage site was fused to the C-terminus of cystatin C, and the protein was well expressed in Escherichia coli after optimization. Then, two different systems were used to obtain no-tag cystatin C: a traditional nickel (Ni)-column system and a subtly Ni magnetic bead system. The column system was more commonly used, and the magnetic bead system was more convenient. Cystatin C (purity > 97%) was successfully obtained, and the yields in both the systems were higher than those in previous studies. Further, the proper folding status and bioactivity of recombinant cystatin C were confirmed using the papain inhibition assay, dynamic light scattering, and circular dichroism spectroscopy.
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