refolding process of cysteine-rich proteins: chitinase as a model

background: recombinant proteins overexpressed in e. coli are usually deposited in inclusion bodies. cysteines in the protein contribute to this process. inter- and intra- molecular disulfide bonds in chitinase, a cysteine-rich protein, cause aggregation when the recombinant protein is overexpressed in e. coli. hence, aggregated proteins should be solubilized and allowed to refold to obtain native- or correctly- folded recombinant proteins. methods: dilution method that allows refolding of recombinant proteins, especially at high protein concentrations, is to slowly add the soluble protein to refolding buffer. for this purpose: first, the inclusion bodies containing insoluble proteins were purified; second, the aggregated proteins were solubilized; finally, the soluble proteins were refolded using glutathione redox system, guanidinium chloride, dithiothreitol, sucrose, and glycerol, simultaneously. results: after protein solubilization and refolding, sds-page showed a 32 kda band that was recognized by an anti-chitin antibody on western blots. conclusion: by this method, cysteine-rich proteins from e. coli inclusion bodies can be solubilized and correctly folded into active proteins.