[Superproduction of Bacillus intermedius 7P ribonuclease (binase) in Escherichia coli]

We have reported previously about the cloning of the binase gene in E. coli. In this work, using an original approach named "homolog gene recombination" method (HGR), vectors for binase expression in E. coli have been constructed. Transcription of the binase gene have been directed through either tac-promoter or PR-promoter of bacteriophage lambda under the control of temperature-sensitive CI857 repressor. The last promoter gave the maximum yield of binase, up to 100 mg of protein per litre of heat-induced bacterial culture. The location of the transcription terminator at the 3' terminus of the binase gene raised the expression approximately two times more. A chromatographic method have been developed and applied for the control of binase accumulation in growth medium without measuring the ribonuclease activity.