Improving Surfactin Production in Bacillus subtilis 168 by Metabolic Engineering

Area of Science:

• Microbiology
• Biotechnology
• Metabolic Engineering

Background:

• Surfactin is a potent biosurfactant with diverse industrial applications.
• Wild-type Bacillus subtilis 168 lacks the ability to synthesize surfactin.
• Genetic engineering strategies are needed to establish efficient surfactin production.

Purpose of the Study:

• To engineer Bacillus subtilis 168 for enhanced surfactin production.
• To investigate the roles of specific genes in surfactin biosynthesis and transport.
• To optimize fermentation conditions for high-yield surfactin production.

Main Methods:

• Overexpression of the phosphopantetheinyl transferase (PPTase) gene sfp*.
• Gene knockout strategies targeting ppsD, yvkC, and codY.
• Overexpression of surfactin transporter genes (yfiS, yerP, ycxA).
• Fermentation optimization using fed-batch culture in a 5 L fermenter.

Main Results:

• Restored surfactin synthesis in B. subtilis 168 via sfp* overexpression (747.5 mg/L).
• Identified YfiS as a potential surfactin efflux transporter.
• Achieved a significantly enhanced surfactin titre (1601.8 mg/L) by knocking out the codY gene.
• Reached a maximum surfactin titre of 3.89 g/L with a yield of 0.63 g/g DCW.

Conclusions:

• Successful engineering of B. subtilis 168 for high-yield surfactin production.
• CodY is a key negative regulator of surfactin synthesis.
• The engineered strain provides a robust platform for industrial surfactin manufacturing.