Increasing the stability of sacB transcript improves levansucrase production in Bacillus subtilis.

Aims: To develop a strategy to increase the stability of transcripts of structural genes expressed under the control of sacR, the leader region of Bacillus subtilis levansucrase gene. Methods and Results: Insertion of Shine Dalgarno like sequences in the 5'-untranslated sacR region controlling the expression of sacB. Depending on the number of stabilizing sequences inserted and the position of these sequences with respect to the translation start codon, it was observed that the mRNA stability and the final protein production could be increased or decreased. Conclusions: This mRNA stabilization can be used to increase exocellular protein production in the degU32 (Hy) mutant. Significance and Impact of the Study: This approach can be applied to the expression of heterologous genes of biotechnological interest

Autogenous modulation of the Bacillus subtilis sacB-levB-yveA levansucrase operon by levB transcript.

Silencing of levB, the second structural gene of the tricistronic levansucrase operon encoding the endolevanase LevB, decreases the level of levansucrase expression. Conversely, independent expression of levB greatly stimulates operon expression in Bacillus subtilis. This autogenous effect is mediated by the levB transcript, which carries an internal sequence (5'-AAAGCAGGCAA-3') involved in the enhancing effect. In vitro, the levB transcript displays an affinity to the N-terminal fragment of SacY (KD 0.2 µM), the regulatory protein that prevents transcription termination of levansucrase operon. This positive feed back loop leads to an increase in the operon expression when B. subtilis is growing in the presence of high sucrose concentrations. Under these conditions, extracellular levan synthesized by the fructosyl polymerase activity of levansucrase can be degraded mainly into levanbiose by the action of LevB. Levanbiose is neither taken up nor metabolized by the bacteria. This work modifies the present view of the status of levansucrase in B. subtilis physiology

L'opéron lévane saccharase de Bacillus subtilis (régulation transcriptionnelle et post-transcriptionnelle)

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