P141: Mechanosensitive channels as potentially versatile exporters

 Corynebacterium glutamicum is used worldwide in the industrial fermentative production of glutamate. However, the mechanism underlying glutamate secretion by this bacterium remains unclear. A mutation in NCgl1221—a gene encoding a mechanosensitive channel—is reported to cause spontaneous glutamate secretion. In this study, we used an electrophysiological technique (patch clamping) to analyze the functions of this channel in Bacillus subtilis cells expressing NCgl1221. Additionally, we aimed to create a versatile exporter that exploits the properties of the mechanosensitive channel.

 B. subtilis possesses 4 genes (mscL, ykuT, yhdY, and yfkC) that encode mechanosensitive channel homologs. When we applied turgor pressure to giant provacuoles prepared from the mscL, ykuT, yhdY, and yfkC quadruple-disruptant strain of B. subtilis expressing NCgl1221, we observed a pressure-dependent current in membrane tension of 2 mN/m or more. This current appeared to represent the movement of potassium ions from inside to outside the provacuole. Furthermore we found that glutamic acid and aspartic acid were transported across the cytoplasmic membrane through the NCgl1221channel in by passive diffusion. Therefore, we considered using this channel as a versatile exporter.

 A gain-of-function mutation in NCgl1221 was expressed in a phenylalanine producer (Escherichia coliAJ12741). This strain displayed remarkably high phenylalanine productivity compared with a control strain. These data suggest that inducing the mechanosensitive channel as a versatile exporter could be effective in improving fermentation yield. We believe that the findings of this study could make a significant contribution to developments in exporter engineering (ExE).