Improving the production of the micafungin precursor FR901379 in an industrial production strain

Abstract

Background Micafungin is an echinocandin-type antifungal agent used for the clinical treatment of invasive fungal infections. It is semisynthesized from the sulfonated lipohexapeptide FR901379, a nonribosomal peptide produced by the filamentous fungus Coleophoma empetri. However, the low fermentation efficiency of FR901379 increases the cost of micafungin production and hinders its widespread clinical application. Results Here, a highly efficient FR901379-producing strain was constructed via systems metabolic engineering in C. empetri MEFC09. First, the biosynthesis pathway of FR901379 was optimized by overexpressing the rate-limiting enzymes cytochrome P450 McfF and McfH, which successfully eliminated the accumulation of unwanted byproducts and increased the production of FR901379. Then, the functions of putative self-resistance genes encoding β-1,3-glucan synthase were evaluated in vivo. The deletion of CEfks1 affected growth and resulted in more spherical cells. Additionally, the transcriptional activator McfJ for the regulation of FR901379 biosynthesis was identified and applied in metabolic engineering. Overexpressing mcfJ markedly increased the production of FR901379 from 0.3 g/L to 1.3 g/L. Finally, the engineered strain coexpressing mcfJ, mcfF, and mcfH was constructed for additive effects, and the FR901379 titer reached 4.0 g/L under fed-batch conditions in a 5 L bioreactor. Conclusions This study represents a significant improvement for the production of FR901379 and provides guidance for the establishment of efficient fungal cell factories for other echinocandins.

Industrial Production of Antibiotics in Fungi: Current State, Deciphering the Molecular Basis of Classical Strain Improvement and Increasing the Production of High-Yielding Strains by the Addition of Low-Molecular Weight Inducers

“…Lipopeptide FR901379 is obtained as a result of the natural fermentation of the improved strains of the ascomycete Coleophoma empetri [254,255]. Recently, the industrial producer C. empetri MEFC009, using a genetic engineering approach, was able to further increase the yield of FR901379 [182,256].…”

Insight into advances for the biosynthetic progress of fermented echinocandins of antifungals

“…In the field of fermentation, the augmentation of production yields of pneumocandin B 0 , echinocandin B, and FR901379 has been meticulously pursued through the optimization of carbon–nitrogen sources and cultivation conditions. As a result, the pinnacle of achievement has been marked by yield reaching about 2530, 3150, and 4000 mg/L for pneumocandin B 0 , echinocandin B, and FR901379, respectively (Men et al., 2023 ; Niu et al., 2020 ; Yuan et al., 2019 ).…”

“…On the other hand, GLhyp / ecdJ / mcfJ lacks apparent structural domains, but mcfJ acts as a transcriptional activator such that deletion of mcfJ in C. empetri leads to the loss of FR901379 production accompanied with nearly all the involved genes becoming silent (Wei et al., 2022 ). Conversely, overexpression of mcfJ can increase FR901379 production (Men et al., 2023 ).…”

“…These novel echinocandins exhibit distinct antifungal activities (Wei et al., 2022 ). In addition to these advancements, a gene overexpression strategy has been implemented in C. empetri to obtain the high‐yielding strain of FR901379; overexpression of mcfJ , mcfF , and mcfH genes led to a substantial increase in FR901379 production, reaching a maximum titre about to 4000 mg/L, representing a 13‐fold increase (Men et al., 2023 ). Overall, these mutants have significantly increased the yield and purity of echinocandins, facilitating the generation of new products.…”