An Engineered Microbial Consortium Provides Precursors for Fengycin Production by Bacillus subtilis

Abstract

Fengycin has great potential for applications in biological control because of its biosafety and degradability. In this study, the addition of exogenous precursors increased fengycin production by Bacillus subtilis. Corynebacterium glutamicum was engineered to produce high levels of precursors (Thr, Pro, Val, and Ile) to promote the biosynthesis of fengycin. Furthermore, recombinant C. glutamicum and Yarrowia lipolytica providing amino acid and fatty acid precursors were co-cultured to improve fengycin production by B. subtilis in a threestrain artificial consortium, in which fengycin production was 2100 mg• L −1 . In addition, fengycin production by the consortium in a 5 L bioreactor reached 3290 mg•L −1 . Fengycin had a significant antifungal effect on Rhizoctonia solani, which illustrates its potential as a food preservative. Taken together, this work provides a new strategy for improving fengycin production by a microbial consortium and metabolic engineering.

Strategies for improving fengycin production: a review

“…Furthermore, the carbon to nitrogen ratio is also crucial, with different ratios favoring different microbial strains. The optimal carbon to nitrogen ratio for lipopeptide production in B. amyloliquefaciens MEP218 is 10:1 using glucose and NH 4 NO 3 [ 60 ], while Wei et al [ 61 ] achieved the highest fengycin titer of 1220 mg/L with a carbon to nitrogen ratio of approximately 8. Since different strains may have different nitrogen source requirements, it is advisable to perform a case-by-case analysis of the selection.…”

“…It has been demonstrated that the addition of fatty acids (including myristic acid, pentadecanoic acid, heptadecanoic acid, nonadecanoic acid, and C16 fatty acid) [ 14 , 61 ] and amino acids (such as glutamic acid, asparagine, serine, alanine, lysine, and ornithine) [ 54 , 56 , 66 , 67 ] significantly enhanced the production of fengycin, in which the addition of alkanoic acid could up-regulate the transcription levels of synthetic genes fenA , srfAA , ituD , and fatty acid metabolism-related genes fabI and fadB [ 14 ], while addition of 10 g/L glutamic acid enhanced fengycin production mainly by up-regulating the expression of membrane transport systems [ 54 ]. Nevertheless, the addition of the identical amino acids may also result in different effects on fengycin production in different strains [ 54 , 61 , 66 ]. This variation may be attributed to differences in the distribution of metabolic fluxes in different strains, which in turn result in varying requirements for precursor amino acids.…”

“…To meet with the market demand for fengycin, numerous studies have been devoted to increasing the production of fengycin through metabolic engineering approaches, including enhancement of precursor supply [ 46 , 48 , 54 , 56 , 57 , 61 , 73 – 75 ], application of regulatory factors [ 8 , 45 , 64 , 75 – 84 ], promoter engineering [ 45 , 48 , 67 , 75 , 79 , 85 – 87 ], and application of genome-engineering (genome shuffling [ 20 , 88 ] and genome-scale metabolic network model [ 74 ]).…”

“…Corynebacterium glutamicum is an industrial strain capable of producing various amino acids, making it an excellent choice for providing amino acids in a co-culture system [ 56 ]. Several studies have shown that co-cultivation of fengycin-producing strains with a series of C. glutamicum , which have high production of proline, serine, threonine, valine and isoleucine, resulted in significantly higher fengycin production due to the provision of sufficient amino acid precursors, compared with pure cultures [ 56 , 57 , 61 , 73 ]. However, the introduction of an engineered Saccharomyces cerevisiae , which can hydrolyze the starch from food waste to provide carbon source, into the three-strain artificial consortia for fengycin production resulted in a decrease in lipopeptide yields [ 57 ].…”

“…This may be due to substrate competition and energy allocation in the co-culture system of the four strains. It’s worth noting that inoculation time and ratio of different strains, as well as culture medium used for co-cultivation are also significantly important for lipopeptides production [ 56 , 61 ]. Therefore, although microbial co-culture has shown great potential for fengycin production, designing and optimizing microbial communities remains a major challenge.…”