Protein folding studies in vivo 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-beta-galactosidase, IPIII-beta-globin, and IPIII-beta-globin-beta-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 beta-galactosidase. Phage inject IPIII-beta-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 beta-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.