We have developed an approach to produce insoluble hydrophobic proteins in stable soluble forms. The proposed system is a fused protein construct, where a target protein is fused to the carrier. The carrier is GrAD protein, it is apical domain of thermophillic minichaperone GroEL. The Met-less version of GroEL apical domain has been made for further use of CNBr chemical cleavage. Studied proteins included hydrophobic insoluble proteins fused to the C- terminus of GrAD as well as its N-terminus. The fusions were expressed at various temperatures and inclusion bodies were harvested. All fusion proteins were dissolved in urea solution and successfully renaturated in stable soluble forms. Their stability and target protein functionality were tested. This approach can be used to solubilize and stabilize hydrophobic proteins for their research by different means.
GroEL, protein stabilization, hydrophobic proteins, chaperones, minichaperone, GroEL
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