Cloning, expression and characterization of the Jiangella aurantiaca encapsulin system

Encapsulins are natural, bacterial-derived, protein-structured capsules with specific cargo proteins. In this study, the peroxidase-encapsulin system of Jiangella aurantiaca 8K307 was characterized. In addition, the effects of encapsulin on oligo-1,6-α-glucosidase were investigated by adding the cargo loading peptide (CLP) belonging to Dyp to the oligo-1,6-α-glucosidase of Anoxybacillus ayderensis A9, which is unrelated to encapsulin. Encapsulin, Dyp, oligo-1,6-α-glucosidase and CLP-labeled oligo-1,6-α-glucosidase genes were produced recombinantly and obtained in pure form after gene expression. The 24 nm capsule structure of encapsulin was determined by TEM and DLS analyses. Physiological conditions under which the capsule form was stable were determined. In vitro encapsulation of encapsulin was achieved with Dyp and CLP labeled oligo-1,6-α-glucosidase. Encapsulations were detected by zymogram and Native PAGE analyses. Optimum pH, optimum temperature and kinetic parameters of encapsulinated and free Dyp, encapsulinated and free oligo-1,6-α-glucosidase, and oligo-1,6-α-glucosidase without CLP label were determined. The effects of surfactants, metal chelators, reducing agents, organic solvents, and metals on Dyp, oligo-1,6-α-glucosidase, and CLP-tagged oligo-1,6-α-glucosidase were tested. As a result, the protective effects of encapsulin on the specific cargo protein Dyp and the unrelated cargo protein oligo-1,6-α-glucosidase were characterized in detail