A tightly regulated system for overproduction of bacteriophage T4 lysozyme in Escherichia coli

Bacteriophage T4 lysozyme has been purified using the Ni-chelate affinity chromatography technique from overexpressing Escherichia coli cells by fusion to an N-terminal 6x His tail. Regulation of the lysozyme gene expression has been found to be critical during growth phase of the bacteria by comparing different plasmid constructions. Whereas a tac-promoter fusion construct alone did not lead to efficient production of T4 lysozyme because of early cell lysis, an improved repressor sequence and co-overproduction of the tac repressor resulted in high-level synthesis of the foreign protein after IPTG induction. Purification of the fusion protein from autolyzed crude cell extracts is possible in a simple one-step procedure.