Enhancement of ε-poly-lysine production in ε-poly-lysine-tolerant Streptomyces sp. by genome shuffling

Abstract

ε-Poly-L-lysine (ε-PL) has been widely used as food additive. However, the self-inhibition of ε-PL on cell growth limits the accumulation of ε-PL in the wild-type strain. Here, we screened ε-PL-tolerant strain of Streptomyces sp. with higher ε-PL productivity by genome shuffling and studied the mechanism for the improvement. The initial mutant library was constructed by diethyl sulfate mutagenesis. After four rounds of protoplast fusion, a shuffled strain F4-22 with 3.11 g/L ε-PL productivity in shake flask, 1.81-fold in comparison with that of parent strain, was obtained. The higher aspartokinase activity was induced in F4-22 whereas no obvious changes have been found in ε-PL synthetic and degrading enzymes which indicated that the upstream reregulation of the precursor lysine synthesis rather than lysine polymerization or ε-PL degradation in shuffled strain accounted for the higher productivity. The activities of key enzymes in the central metabolic pathway were also enhanced in F4-22 which resulted in increased vigor of the strain and in delayed strain lysis during fermentation. These improved properties of shuffled strain led to the success of combining general two-stage fermentation into one-stage one in 5-L bioreactor with 32.7 % more ε-PL production than that of parent strain. The strategy used in this study provided a novel strain breeding approach of producers which suffered from ε-PL-like self-inhibition of the metabolites.

Identification of ε-Poly-L-lysine as an Antimicrobial Product from an Epichloë Endophyte and Isolation of Fungal ε-PL Synthetase Gene

“…Since ε-PL has attracted attention as an antimicrobial agent with a wide inhibitory spectrum against Gram-positive and negative bacteria, fungi, yeasts, and phages [25], several strategies to improve the production of ε-PL have been conducted; for example, modifying culture conditions (supplementation with glucose or other metabolic precursors, modifying pH, etc.) [27,34], inducing double antibiotic-resistant mutations [35] and genome shuffling [36]. The present study suggests that the gene overexpression strategy under the control of the TEF promoter is one of the excellent strategies for the high production of ε-PL.…”

Epsilon-poly-L-lysine: Recent Advances in Biomanufacturing and Applications

“…More recently, similar work facilitated the integration of traits from S. albulus and B. subtilis enabling ɛ-PL production 256.1% higher than that of the parent strain ( Li et al, 2018 ). Additionally, since the auto-inhibition caused by the accumulation of ɛ-PL may limit cell growth and ɛ-PL biosynthesis, genome shuffling has been also used to rapidly increase the ɛ-PL tolerance and production ( Zhou et al, 2015 ).…”

The genome and transcriptome of Lactococcus lactis ssp. lactis F44 and G423: Insights into adaptation to the acidic environment

“…Contrary to laborious classical mutagenesis and directional evolution, which require massive relative genetic data and metabolic network information, genome shuffling (a recursive protoplast) has been widely applied to studies on stress tolerance (Zhang et al, 2002;Gerando et al, 2016;Zhu et al, 2016) and production enhancement (Wang et al, 2014;Zhou et al, 2015;Yin et al, 2016). This technology has been used to enhance the nisin productivity (Zhang et al, 2014) and acid tolerance of Lc.…”