Altered temperature induction sensitivity of the lambda p sub(R)/cI857 system for controlled gene E expression in Escherichia coli

Cell lysis of Gram-negative bacteria can be efficiently achieved by expression of the cloned lysis gene E of bacteriophage PhiX174. Gene E expression is tightly controlled by the rightward lambda p sub(R) promoter and the temperature-sensitive repressor cI857 on lysis plasmid pAW12. The resulting empty bacterial cell envelopes, called bacterial ghosts, are currently under investigation as candidate vaccines. Expression of gene E is stringently repressed at temperatures up to 30 degree C, whereas gene E expression, and thus cell lysis, is induced at temperatures higher than 30 degree C due to thermal inactivation of the cI857 repressor. As a consequence, the production of ghosts requires that bacteria have to be grown at 28 degree C before the lysis process is induced. In order to reflect the growth temperature of pathogenic bacteria in vivo, it seemed favorable to extend the heat stability of the lambda p sub(R) promoter/cI857 repressor system, allowing pathogens to grow at 37 degree C before induction of lysis. In this study we describe a mutation in the lambda p sub(R) promoter, which allows stringent repression of gene E expression at temperatures up to 36 degree C, but still permits induction of cell lysis at 42 degree C.