Bent promoter of epsilon-toxin gene from Clostridium perfringens : analysis and application to protein expressions.

阶段性a束形成DNA的弯曲结构。我们发现了一种新型的弯曲DNA结构，它可以刺激产气荚膜梭菌毒素启动子下游的基因转录。我们比较了ε -毒素启动子与其他弯曲启动子、α -毒素启动子和铁氧还蛋白启动子基因转录的温度依赖性；Epsilon-toxin、α -toxin和铁氧还蛋白启动子分别在37℃、25℃和较宽的温度范围内表现出高水平的转录。因此，我们构建了一种新的表达载体，利用铁氧还蛋白启动子在产气荚膜荚膜杆菌中高效表达大型梭菌蛋白。我们的表达系统成功地产生了大量大肠杆菌表达系统中表达较差的ColH、NagH、NagJ、NanJ和BgaA。这些蛋白从重组产气荚膜杆菌培养的上清液中得到了足够的纯化量。

Phased A-tracts form a bent structure of DNA. We found a novel type of bent DNA structure, which stimulates the gene transcription, downstream of a Clostridium perfringens epsilon-toxin promoter. When we compared the temperature dependency of gene transcription from epsilon-toxin promoter to those from other bending promoters, alpha-toxin and ferredoxin promoters ; epsilon-toxin, alpha-toxin and ferredoxin promoters exhibit high-level transcription at 37℃, 25℃ and over a wide rangeof temperature, respectively. Thus, we constructed a novel expression vector using ferredoxin promoter to express efficiently large clostridial proteins in C.perfringens. Our expression system was successfully applied to produce large amounts of ColH, NagH, NagJ, NanJ, and BgaA, which are poorly expressed in Escherichia coli expression system. These proteins were purified in sufficient amounts from the supernatants of the recombinant C.perfringens cultures.