A temperature‐sensitive expression system based on the Geobacillus stearothermophilus NRS 2004/3a sgsE surface‐layer gene promoter

The sgsE gene coding for the S‐layer (surface layer) protein in the thermophilic Gram‐positive bacterium Geobacillus stearothermophilus NRS 2004/3a is strongly induced when the culture is shifted from optimal (55 °C) to maximally tolerable growth temperature (67 °C). Here, we investigated the regulation of the sgsE promoter in G. stearothermophilus and tested the function of this promoter in Bacillus subtilis . We used EGFP (enhanced green fluorescent protein) reporter constructs and found that the sgsE promoter has very low basal activity at 28 °C, but is approx. 20‐fold induced by elevated growth temperatures (37 and 45 °C). The promoter confers high expression levels, as EGFP mRNA levels at 45 °C were approx. 120‐fold more abundant than mRNA levels of the cat (chloramphenicol resistance) gene, which was transcribed from a constitutive promoter on the same plasmid. In fluorescence‐microscopic and Western‐blot analysis, the EGFP protein was barely detectable at 28 °C, whereas intermediate and high levels were detected at 37 and 45 °C respectively. The potential to tune expression levels of genes driven by the sgsE promoter in B. subtilis by simple temperature adjustments presents a considerable potential for its future use as high‐yield protein expression system for B. subtilis .