Abstract
The production of pneumocandin B0 is limited by feedback inhibition. Here, low-temperature adaptive laboratory evolution (ALE) was used to improve the production capacity of Glarea lozoyensis by enhancing its membrane permeability. After 50 cycles of ALE, the pneumocandin B0 production of the endpoint strain (ALE50) reached 2131 g/L, which was 32% higher than the starting strain (ALE0). ALE50 showed a changed fatty acid composition of the cell membrane, which-+h increased its permeability by 14%, which in turn increased the secretion ratio threefold. Furthermore, ALE50 showed increased intracellular proline and acetyl-CoA concentrations, superoxide dismutase (SOD), and catalase (CAT) activity, as well as total antioxidant capacity. The slight biomass decrease in ALE50 was accompanied by decreased isocitrate dehydrogenase (ICDH) and glucose-6-phosphate dehydrogenase (G6PDH) activity. Finally, a putative model of the accumulation and secretion of pneumocandin B0 in ALE50 was established. ALE is a promising method to release intracellular feedback inhibition.
Highlights
Pneumocandin B0 is a lipohexapeptide composed of a 10R,12S-dimethylmyristoyl side chain and a hexapeptide core, which is produced by the fungus Glarea lozoyensis (Li et al, 2015)
The aim of this study was to provide a new method based on adaptive laboratory evolution (ALE) for the improvement of the productivity of pneumocandin B0 by G. lozoyensis, but this approach may be useful in other fungal species that are used for the fermentation of intracellular products
The starting strain ALE0 of G. lozoyensis was cultivated under low temperature conditions for 50 cycles, and ALE strains were collected at the end of each 10 cycles to monitor the glucose consumption and dry cell weight (DCW) during ALE (Figure 1A), which was recognized as first indicators of the success or failure of ALE
Summary
Pneumocandin B0 is a lipohexapeptide composed of a 10R,12S-dimethylmyristoyl side chain and a hexapeptide core, which is produced by the fungus Glarea lozoyensis (Li et al, 2015). Caspofungin, a semi-synthetic derivative of pneumocandin B0, is the first member of the echinocandins family approved of the treatment of fungal infection by the FDA. Genomic and metabolomic approaches have already been used to improve pneumocandin biosynthesis (Youssar et al, 2012; Chen et al, 2013, 2015; Song et al, 2018). Pneumocandin B0 is mainly biosynthesized and accumulated in the mycelia (Balkovec et al, 2014). It is challenging to achieve high pneumocandin B0 productivity inside the mycelia due to feedback inhibition.
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