Mutation analysis of pertussis toxin promoter.
The improving of the expression efficiency of a pertussis toxin (PT) promoter was believed to be a critical issue for the production of PT in acellular vaccine development. In this study, we have isolated a PT promoter region from the genome of a pertussis strain ATCC 9340. Based on the promoter sequence, a series of mutant PT promoters have been generated and subjected to in vitro gel shift analysis and in vivo reporter beta-galactosidase activity study. As compared with the wild type promoter, the mutation of the ribosome binding sequence or -10 element, to the respective consensus sequence derived from strong bacterial promoters, resulted in an enhancement of its interaction with two cellular proteins, and a slightly higher beta-galactosidase activity (1.3 fold). Whereas, the change of either upstream inverse repeats or 20-bp direct repeats to a certain complete repeat significantly promoted the formation of another DNA-protein-complex, and exhibited an 1.8 fold beta-galactosidase activity. These findings would have provided a mutation target for making a more efficient PT-production pertussis strain.
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Division of Biologics Quality Control, National Institute of Preventive Medicine, Taipei, Taiwan, ROC.
Chiu
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SC
Lu
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D001483:Base Sequence; D008969:Molecular Sequence Data; D009154:Mutation; D037342:Pertussis Toxin; D011401:Promoter Regions, Genetic; D010566:Virulence Factors, Bordetella / Q000096:biosynthesis / Q000235:genetics*
D010566:Virulence Factors, Bordetella; D037342:Pertussis Toxin