Construction and description of a constitutive plipastatin mono-producing Bacillus subtilis

Abstract

Background Fengycin and plipastatin are potent Bacillus antimicrobial lipopeptide with the prospect to replace conventional antifungal chemicals for controlling plant pathogens. However, the application of these lipopeptides has so far been investigated in a few cases, principally because of their yield in low concentration and unknown regulation of biosynthesis pathways. B. subtilis synthesizes plipastatin by a non-ribosomal peptide synthetase encoded by the ppsABCDE operon. In this study, B. subtilis 3NA (a non-sporulation strain) was engineered to construct an efficient plipastatin mono-producer strain and to gain more insights about bottlenecks of plipastatin production.Results The first step toward the construction of a plipastatin mono-producer was repairing the defective sfp gene in strain 3NA to construct strain BMV9. Production of plipastatin was doubled after repairing the gene expression of the pleiotropic regulator degQ (strain BMV10) compared to BMV9. Moreover, substitution of the plipastatin promoter (Ppps) against the strong constitutive promoter Pveg led to approximately fivefold enhancement of plipastatin production in BMV11 compared to BMV9. Intriguingly, combination of both repaired degQ expression and promoter exchange (Ppps::Pveg) did not increase the plipastatin yield. Surprisingly, deletion of surfactin (srfAA-AD) operon by the retaining the functional comS gene, which is located within srfAB, resulted in the loss of plipastatin production in BMV9 and significantly decreased the plipastatin production of BMV11. We also observed that supplementation of ornithine as a precursor for plipastatin formation caused higher production of plipastatin in mono-producer strains, albeit with a modified pattern of plipastatin composition.Conclusions This study provides evidence that degQ stimulates the native plipastatin production. Surprisingly, it seems that surfactin synthetase or one of its subunits positively regulates plipastatin production by an unknown mechanism. Furthermore, as another conclusion of this study, results point towards ornithine provision being a bottleneck for plipastatin production. Therefore, targeting the ornithine metabolic flux might be a promising strategy to further investigate and enhance plipastatin production.

Exploration of surfactin production by newly isolated Bacillus and Lysinibacillus strains from food-related sources

“…In this way, three strains were found to produce both surfactin and fengycin (EF1-A2, EFSO2-04, POC4-2). However, fengycin is generally produced only in small amounts, which makes the detection in some cases difficult (Vahidinasab et al 2020). Moreover, five strains (E2-02, EFS1-2, E2-03, IRB4-02) were found for the production of surfactin and iturin, with ES1-02 (Fig.…”

Screening, Production, and Characterization of Antifungal Secondary Metabolites From Bacillus Velezensis Alo Na L4 of Aloe Vera

“…The lipopeptides [63,64] in addition to several modified small peptides as well as proteins [65,66]. Bacillus velezensis is a Grampositive, rod-shaped bacterium that forms spores, and has long been recognized as one of the most important bacterium that is potentially promising to generate many antimicrobial compounds [67,68] with potential applications in the various fields.…”