The production of recombinant penicillin acylase (PAC) in Escherichia coli was optimized in this study. The effect of using a selection of host/vector systems as well as varying culture conditions on the production of PAC was investigated. The production of PAC based on the use of the native pac promoter was inefficient and could be significantly improved by using the strong trc promoter for regulation of pac expression. A mutant strain MDDeltaP7 was shown to be a suitable host for the production of PAC since the efficiency of both pac translation and posttranslational processing for MDDeltaP7 was significantly higher than that for the parent strain HBPAC101. However, the accumulation of inclusion bodies tended to limit the production of PAC as pac transcriptional and translational efficiency was increased. It has been demonstrated that, in addition to the increase in pac transcriptional and translational efficiency, the protein synthesis flux throughout pac expression steps should be balanced for enhancing the production of PAC in E. coli. With the optimization of the host/vector system and culture conditions, culture performance for the production of recombinant PAC was greatly improved. Process bottlenecks limiting the production of PAC were also discussed.

Lin, WJ Kuo, BY Chou, CP