Optimization of staphylokinase production inBacillus subtilis using inducible and constitutive promoters

Staphylokinase (SAK) was produced inB. subtillis using two different promoter systems,i.e. the P43 andsacB promoters. To maximize SAK expression inB. subtilis, fermentation control strategies for each promoter were examined. SAK, under P43, a vegetative promoter transcribed mainly by σ^B containing RNA polymerase, was overexpressed at low dissolved oxygen (D.O.) levels, suggesting that thesigB operon is somewhat affected by the energy charge of the cells. The expression of SAK at the 10% D.O. level was three times higher than that at the 50% D.O. level. In the case ofsacB, a sucrose-inducible promoter, sucrose feeding was used to control the induction period and induction strength. Since sucrose is hydrolyzed by two sucrose hydrolyzing enzymes in the cell and culture broth, the control strategy was based on replenishing the loss of sucrose in the culture. With continuous feeding of sucrose, WB700 (pSAKBQ), which contains the SAK gene undersacB promoter, yieldedca. 35% more SAK than the batch culture. These results present efficient promoter-dependent control strategies inB. subtilis host system for foreign protein expression.