PROTEIN FOLDING STUDIES INVIVO WITH A BACTERIOPHAGE-T4 EXPRESSION-PACKAGING-PROCESSING VECTOR THAT DELIVERS ENCAPSIDATED FUSION PROTEINS INTO BACTERIA

A cloned phage T4 gene which expresses the nonessential capsid scaffold protein IPIII was modified to permit construction and packaging of protein fusions within the capsid. IPIII deletion phage packaged IPIII-β-galactosidase, IPIII-β-globin, and IPIII-β-globin-β-galactosidase fusion proteins; the latter protein fusion was specifically processed by the T4 gene 21 head morphogenetic proteinase in vivo at a consensus leu(ile)-P1-glu cleavage site to regenerate β-galactosidase. Phage inject IPIII-β-galactosidase protein into bacteria, but less activity is recovered in infections of Escherichia coli dnaK or groEL mutants, suggesting that these host molecular chaperones are required for β-galactosidase intracellular folding. This expression-packaging-processing (EPP) vector directs protein fusions into capsids for easy detection and purification and permits study of protein delivery and folding in bacteria. © 1993 Academic Press. All rights reserved.