Streptomyces Albulus Research Articles

Metabolic perturbation of Streptomyces albulus by introducing NADP-dependent glyceraldehyde 3-phosphate dehydrogenase.

The available resources of Streptomyces represent a valuable repository of bioactive natural products that warrant exploration. Streptomyces albulus is primarily utilized in the industrial synthesis of ε-poly-L-lysine (ε-PL). In this study, the NADP-dependent glyceraldehyde 3-phosphate dehydrogenase (GapN) from Streptococcus mutans was heterologously expressed in S. albulus CICC11022, leading to elevated intracellular NADPH levels and reduced NADH and ATP concentrations. The resulting perturbation of S. albulus metabolism was comprehensively analyzed using transcriptomic and metabolomic methodologies. A decrease in production of ε-PL was observed. The expression of gapN significantly impacted on 23 gene clusters responsible for the biosynthesis of secondary metabolites. A comprehensive analysis revealed a total of 21 metabolites exhibiting elevated levels both intracellularly and extracellularly in the gapN expressing strain compared to those in the control strain. These findings underscore the potential of S. albulus to generate diverse bioactive natural products, thus offering valuable insights for the utilization of known Streptomyces resources through genetic manipulation.

Identification and Combinatorial Overexpression of Key Genes for Enhancing ε-Poly-L-lysine Biosynthesis in Streptomyces albulus

ε-Poly-L-lysine (ε-PL) is a natural and safe food preservative mainly produced by the aerobic, filamentous bacterium Streptomyces albulus. Therefore, it is crucial to breed superior ε-PL-producing strains to enhance fermentation efficiency to reduce production costs. Metabolic engineering is an effective measure for strain modification, but there are few reports on key genes for ε-PL biosynthesis. In this study, metabolic flux analysis was employed to identify potential key genes in ε-PL biosynthesis in S. albulus WG-608. A total of six potential key genes were identified. Three effective key genes (ppc, pyc and pls) were identified for the first time in ε-PL biosynthesis through overexpression experiments. It also presents the first demonstration of the promoting effects of ppc and pyc on ε-PL biosynthesis. Three genes were then co-expressed in S. albulus WG-608 to obtain OE-ppc-pyc-pls, which exhibited an 11.4% increase in ε-PL production compared to S. albulus WG-608, with a 25.5% increase in specific ε-PL production. Finally, the metabolic flux analysis of OE-ppc-pyc-pls compared to S. albulus WG-608 demonstrated that OE-ppc-pyc-pls successfully altered the metabolic flux as expected. This study not only provides a theoretical basis for the metabolic engineering of ε-PL-producing strains but also provides an effective approach for the metabolic engineering of other metabolites.

Dynamic Responses of Streptomyces albulus QLU58 and Its Acid-Tolerant Derivatives to the Autoacidification in ε-Poly-l-Lysine Production

Streptomyces albulus is a kind of safety bacteria that is used to produce a natural food preservative named ε-poly-l-lysine (ε-PL). Environmental autoacidification (the pH declined from 6.8 to approximately 3.0) inevitably occurred in ε-PL biosynthesis by S. albulus. In this study, the dynamic responses of S. albulus QLU58 and its acid-tolerant mutants to autoacidification were investigated at the physiological and transcriptional levels. The results showed that cell growth, ε-PL production, cell respiratory activity, and intracellular pH (pHi) homeostasis were disturbed by autoacidification. In the initial autoacidification stage (before 24 h), the acid tolerance of S. albulus was effectively improved by increasing the intracellular ATP and related amino acids contents and the H+-ATPase activity, regulating the membrane fatty acids composition, and maintaining the pHi at about 7.7. However, as the autoacidification degree deepened (after 24 h), the metabolic activities decreased and negative cell growth appeared, which weakened the acid tolerance and caused the pHi to decline to about 6.5. Additionally, the acid-tolerant mutants exhibited better performances during autoacidification, which was also confirmed by the related genes’ improved transcription levels. These results provide references for the analysis of progressive environmental modification in ε-PL production.

Engineering Streptomyces albulus to enhance ε-poly-L-lysine production by introducing a polyphosphate kinase-mediated ATP regeneration system

Backgroundε-Poly-L-lysine (ε-PL) is a natural and safe food preservative that is mainly produced by filamentous and aerobic bacteria Streptomyces albulus. During ε-PL biosynthesis, a large amount of ATP is used for the polymerization of L-lysine. A shortage of intracellular ATP is one of the major factors limiting the increase in ε-PL production. In previous studies, researchers have mainly tried to increase the oxygen supply to enhance intracellular ATP levels to improve ε-PL production, which can be achieved through the use of two-stage dissolved oxygen control, oxygen carriers, heterologous expression of hemoglobin, and supplementation with auxiliary energy substrates. However, the enhancement of the intracellular ATP supply by constructing an ATP regeneration system has not yet been considered.ResultsIn this study, a polyphosphate kinase (PPK)-mediated ATP regeneration system was developed and introduced into S. albulus to successfully improve ε-PL production. First, polyP:AMP phosphotransferase (PAP) from Acinetobacter johnsonii was selected for catalyzing the conversion of AMP into ADP through an in vivo test. Moreover, three PPKs from different microbes were compared by in vitro and in vivo studies with respect to catalytic activity and polyphosphate (polyP) preference, and PPK2Bcg from Corynebacterium glutamicum was used for catalyzing the conversion of ADP into ATP. As a result, a recombinant strain PL05 carrying coexpressed pap and ppk2Bcg for catalyzing the conversion of AMP into ATP was constructed. ε-PL production of 2.34 g/L was achieved in shake-flask fermentation, which was an increase of 21.24% compared with S. albulus WG608; intracellular ATP was also increased by 71.56%. In addition, we attempted to develop a dynamic ATP regulation route, but the result was not as expected. Finally, the conditions of polyP6 addition were optimized in batch and fed-batch fermentations, and the maximum ε-PL production of strain PL05 in a 5-L fermenter was 59.25 g/L by fed-batch fermentation, which is the highest ε-PL production reported in genetically engineered strains.ConclusionsIn this study, we proposed and developed a PPK-mediated ATP regeneration system in S. albulus for the first time and significantly enhanced ε-PL production. The study provides an efficient approach to improve the production of not only ε-PL but also other ATP-driven metabolites.

Understanding the Streptomyces albulus response to low-pH stress at the interface of physiology and transcriptomics.

Streptomyces albulus is a well-established cell factory for ε-poly-L-lysine (ε-PL) production. It has been reported that ε-PL biosynthesis is strictly regulated by pH and that ε-PL can accumulate at approximately pH 4.0, which is outside of the general pH range for natural product production by Streptomyces species. However, how S. albulus responds to low pH is not clear. In this study, we attempted to explore the response of S. albulus to low-pH stress at the physiological and global gene transcription levels. At the physiological level, S. albulus maintained intracellular pH homeostasis at ~pH 7.5, increased the unsaturated fatty acid ratio, extended the fatty acid chain length, enhanced ATP accumulation, increased H+-ATPase activity, and accumulated the basic amino acids L-lysine and L-arginine. At the global gene transcription level, carbohydrate metabolism, oxidative phosphorylation, macromolecule protection and repair, and the acid tolerance system were found to be involved in combating low-pH stress. Finally, we preliminarily evaluated the effect of the acid tolerance system and cell membrane fatty acid synthesis on low-pH tolerance via gene manipulation. This work provides new insight into the adaptation mechanism of Streptomyces to low-pH stress and a new opportunity for constructing robust S. albulus strains for ε-PL production. KEY POINTS: • S. albulus consistently remained pH i at ~7.4 regardless of the environmental pH. • S. albulus combats low-pH stress by modulating lipid composition of cell membrane. • Overexpression of cfa in S. albulus could improve low-pH tolerance and ɛ-PL titer.

Induced defense response in soybean to Sclerotinia sclerotiorum using wuyiencin from Streptomyces albulus CK-15.

Sclerotinia sclerotiorum (Lib) de Bary, a destructive fungal pathogen with an extensive host range, causes major economic losses to crop production activities globally. Streptomyces spp. produce secondary metabolites with diverse structures and biological activities, with potential applications in the control of crop disease. This study explored the potential application of wuyiencin, a secondary metabolite of Streptomyces albulus CK-15, to induce defense responses in soybean against S. sclerotiorum. Lesion size was reduced by nearly 60%, in wuyiencin-treated soybean plants compared to plants infected with S. sclerotiorum only, in greenhouse experiments. Wuyiencin induced callose deposition at six hours postinoculation, and increase Reactive-Oxygen-Scavenging enzymes activities, including superoxide dismutase, catalase, and peroxidase activity. Moreover, wuyiencin inoculated before S. sclerotiorum infection significantly increased polyphenol oxidase, phenylalanine ammonia lyase, chitinase, and β-1,3-glucanase activity, suggesting their involvement in soybean defense responses to S. sclerotiorum. Further, qRT-PCR results showed expression levels of the hormone signaling markers CO11, MYC2, PR4, PR1, NPR1, and ERF1, were upregulated in infected leaves treated with wuyiencin.

Influence of Low-Intensity Ultrasound on ε-Polylysine Production: Intracellular ATP and Key Biosynthesis Enzymes during Streptomyces albulus Fermentation.

The effect of low-intensity sonication treatment on cell growth, ε-polylysine (ε-PL) yield and its biological mechanism were investigated, using a 3-L-jar fermenter coupled with an in situ ultrasonic slot with a Streptomyces albulus strain SAR 14-116. Under ultrasonic conditions (28 kHz, 0.37 W cm-2, 60 min), a high biomass of SAR 14-116 and concentration of ε-PL were realized (i.e., they increased by 14.92% and 28.45%, respectively) when compared with a control. Besides this, ultrasonication increased the mycelia viability and intracellular ATP as well as activities of key enzymes involved in the ε-PL biosynthesis pathway, resulting in an improvement in the production of ε-PL. Data on qRT-PCR revealed that ultrasonication also affected the gene expression of key enzymes in the ε-PL biosynthesis pathway, including ε-PL synthetase (PLS). These outcomes provided the basis for understanding the effects of ultrasound-assisted fermentation on the stimulation of metabolite production and fermentation procedure in a fermenter.

A Rhizobacterium, Streptomyces albulus Z1-04-02, Displays Antifungal Activity against Sclerotium Rot in Mungbean.

Sclerotium rot causes damping-off and stem rot in seedlings and mature mungbeans, which negatively impacts cultivation. The use of a rhizobacterium to control soil-borne diseases is an alternative method to the excess use of synthetic fungicides; therefore, this study aims to screen rhizosphere actinobacteria with fungicidal activities against Sclerotium rolfsii, the pathogen that causes sclerotium rot in mungbeans. Primary screening showed that the Streptomyces sp. isolate Z1-04-02 displayed the highest effectiveness against S. rolfsii in dual culture plates, with a percentage inhibition of 74.28%. An assay containing enzymes that degrade cell walls, of the cell-free culture filtrate (CF) of Z1-04-02, showed that the activities of chitinase and β-1,3-glucanase were 0.0209 and 1.0210 U/mL, respectively, which was significantly higher than that of the control (media alone). The cell-free CF of Z1-04-02, incubated at 37 °C and 100 °C, using agar well diffusion, effectively inhibited the growth of S. rolfsii with inhibition percentages of 37.78% and 27.78%, respectively. Solid-phase microextraction (SPME) was applied to trap volatiles released from Z1-04-02 and gas chromatography–mass spectrometry (GC/MS); volatile antifungal compounds were tentatively identified as bicyclic monoterpene (1R)-(-)-myrtenal. The application of the cell-free CF, and the spore suspension of Z1-04-02, showed disease severity indexes (DSIs) of 12.5% and 8.25%, respectively, which were significantly lower than those showing inoculation by S. rolfsii alone. The identification of this strain by morphology, biochemistry tests, and 16s rDNA sequences revealed that Z1-04-02 was Streptomyces albulus. This finding revealed that S. albulus Z1-04-02 displayed diverse fungicidal activities against S. rolfsii, and it has the potential to act as a biological control agent in terms of inhibiting sclerotium rot in mungbeans.

Facile and green synthesis of reduced graphene oxide/loofah sponge for Streptomyces albulus immobilization and ε-poly-l-lysine production

Facile and green fabrication of reduced graphene oxide on loofah sponge (rGOLS) carrier was applied for cell immobilization of ε-Poly-l-lysine (ε-PL) production. Due to surface properties including large specific surface area, high polarity, and low interaction energy, rGOLS-1 was employed as the optimum rGOLS to enhance immobilization of Streptomyces albulus. Compared with raw LS, batch experiments showed rGOLS-1 facilitated superior cell vitality for ε-PL production due to the presence of reduced graphene oxide. In the sequential fed-batch cultivation of Streptomyces albulus using rGOLS-1 with an aerobic plant fibrous-bed bioreactor (rGOLS-1-AFPB), the maximum ε-PL concentration and productivity reached to 39.2 ± 0.63 g/L and 0.48 g/L/h. The cells immobilized in rGOLS-1 with high vitality and ε-PL production efficiency were reused six times over a period of 624 h. This research afforded an effective approach to enhance the fermentation performance of immobilized cells with the design of an advanced immobilization carrier.

Wuyiencin produced by Streptomyces albulus CK-15 displays biocontrol activities against cucumber powdery mildew.

Wuyiencin is a nucleoside antibiotic produced by Streptomyces albulus CK-15. The aim of this study was to determine whether wuyiencin can be used, as a suitable alternative to chemical pesticides, to protect cucumbers (Cucumis sativus L.) from powdery mildew caused by Sphaerotheca fuliginea. Further, the mechanisms underlying the control of cucumber powdery mildew by S. albulus CK-15 were preliminarily elucidated. Wuyiencin solutions of different concentrations were used to treat infected cucumber plants under greenhouse conditions. The results indicated that wuyiencin could significantly reduce powdery mildew disease incidence, with a maximum prevention efficacy of 94·38%. Further, scanning electron micrographs and enzyme assays showed that wuyiencin inhibited S. fuliginea spore growth and elicited the activity of plant systemic resistance-related enzymes. Additionally, real-time quantitative reverse transcription PCR suggested that wuyiencin can activate a salicylic acid-dependent plant defence response. Wuyiencin produced by S. albulus CK-15 possessed antifungal effects and was able to mitigate cucumber powdery mildew disease via antagonistic action. Wuyiencin also induced defence responses in the plants. These results reinforce the biotechnological potential of wuyiencin as both an antagonistic agent and an inducer of plant systemic resistance.

Studying on the antifungal activityof the actinomycete XK1 isolatedfrom growing orange’s soil against pathogenic fungi causing disease in citrus

Streptomyces are considered to be one of the actinomycetes genera having potential in the production of agricultural probiotics because they are safe and have strong antagonism to many bacterial and fungal species that cause plant diseases. In this study, 18 strains of actinomycetes were isolated from growing oranges’ soil in Ha Giang province, of which the actinomycete strain XK1 was considered to be strongly resistant to the fungi Penicillium digitatum and Colletotrichum gloeosporioides causing fruit rot and shedding in citrus. Based on morphological characteristics and 16S rRNA sequence, the strain XK1 was determined to belong to the Streptomyces albulus. On MT2 medium with pH=6, after five days of shaking cultivation at 30°C, the liquid cultures of S. albulus XK1 showed the strongest resisting activity to P. digitatum and C. gloeosporioides. At the same time, the broth cultures of S. albulus XK1 showed inhibitory activity against P. digitatum in oranges. The actinomycete strain S. albulus XK1 was selected base on strong antifungal ability against P. digitatum and C. gloeosporioides and had a potential for application in the production of probiotics used to control fruit rot and shedding in citrus.

Distribution of ε-Poly-l-Lysine Synthetases in Coryneform Bacteria Isolated from Cheese and Human Skin.

ε-Poly-l-lysine is a potent antimicrobial produced through fermentation of Streptomyces and used in many Asian countries as a food preservative. It is synthesized and excreted by a special nonribosomal peptide synthetase (NRPS)-like enzyme called Pls. In this study, we discovered a gene from cheese bacterium Corynebacterium variabile that showed high similarity to the Pls from Streptomyces in terms of domain architecture and gene context. By cloning it into Streptomyces coelicolor with a Streptomyces albulus Pls promoter, we confirmed that its product is indeed ε-poly-l-lysine. A comprehensive sequence analysis suggested that Pls genes are widely spread among coryneform actinobacteria isolated from cheese and human skin; 14 out of 15 Brevibacterium isolates and 10 out of 12 Corynebacterium isolates contain it in their genomes. This finding raises the possibility that ε-poly-l-lysine as a bioactive secondary metabolite might be produced and play a role in the cheese and skin ecosystems.IMPORTANCE Every year, microbial contamination causes billions of tons of food wasted and millions of cases of illness. ε-Poly-l-lysine has potent, wide-spectrum inhibitory activity and is heat stable and biodegradable. It has been approved for food preservation by an increasing number of countries. ε-Poly-l-lysine is produced from soil bacteria of the genus Streptomyces, also producers of various antibiotic drugs and toxins and not considered to be a naturally occurring food component. The frequent finding of pls in cheese and skin bacteria suggests that ε-poly-l-lysine may naturally exist in cheese and on our skin, and ε-poly-l-lysine producers are not limited to filamentous actinobacteria.

Efficacy of bioagents against Pythium deliense Meurs associated with yellowing of black pepper.

Yellowing and wilting of black pepper vines is a serious concern in many black pepper growing tracts where Pythium deliense was recently emerged as a pathogen from the rhizosphere of affected vines, which is proved to be pathogenic by Koch's postulates. As a measure to manage the symptoms, bioagents were evaluated against infection by P. deliense. Among the seven bioagents tested, Trichoderma harzianum and Streptomyces albulus showed 100% inhibition in vitro followed by one Streptomyces sp. and S. rimosus (75.33%). The potential ones were further evaluated under the hydroponic system in vivo by challenge inoculation. No root infection was noticed with T. harzianum and S. albulus inoculation, instead, the inoculated plants showed root regeneration. This suggests the efficiency of these bioagents on plant growth promotion as well as on disease suppression. Biochemical analysis of the hydroponic medium showed an increase in membrane conductivity in all the treatments except in T. harzianum. The release of phenolic compounds into the medium was lowest with T. harzianum indicating the prevention of pathogen invasion. In planta evaluation under greenhouse condition and field evaluation also showed the protective effect of T. harzianum and S. albulus with a reduction in the intensity of yellowing to an extent of 73.1% and 71.2%, respectively. The study revealed that T. harzianum and the actinomycete S. albulus had the potential to prevent the root rot caused by P. deliense.

Facile and Green Synthesis of Reduced Graphene Oxide/Loofah Sponge for Streptomyces Albulus Immobilization and ε-Poly-L-Lysine Production

Facile and Green Synthesis of Reduced Graphene Oxide/Loofah Sponge for Streptomyces Albulus Immobilization and ε-Poly-L-Lysine Production

Metabolomic and genomic profiles of Streptomyces albulus with a higher ε-polylysine production through ARTP mutagenesis

Effects of atmospheric and room temperature plasma (ARTP) mutagenesis on Streptomyces albulus were investigated at cellular and molecular levels to elucidate the intracellular metabolism changes and the mechanism of ε-polylysine (ε-PL) yield enhancement in the mutant strain SAR 14−116 with ARTP treatment. An ε-PL yield increase by 18.46 % was obtained and morphological changes of spores and mycelia were found in SAR 14−116 at a mid-logarithmic phase, compared with its original strain SAB. Kinetic characteristics of ε-PL in the culture broth of SAB and SAR 14−116 were analyzed, showing obvious ε-PL yield difference between these two strains. Untargeted metabolomics analysis showed there were 851 differential metabolites between these two strains. These metabolites were involved with 64 metabolic pathways through KEGG analysis. Genomic analysis showed there were 88 SNPs in SAR 14−116. The ε-PL yield increase in SAR 14−116 might have a relationship with the up-regulated metabolites in the l-lysine synthesis and degradation pathways.

Physiological and Transcriptional Responses of Streptomyces albulus to Acid Stress in the Biosynthesis of ε-Poly-L-lysine.

Streptomyces albulus has commercially been used for the production of ε-poly-L-lysine (ε-PL), a natural food preservative, where acid stress is inevitably encountered in the biosynthesis process. To elucidate the acid tolerance response (ATR), a comparative physiology and transcriptomic analysis of S. albulus M-Z18 at different environmental pH (5.0, 4.0, and 3.0) was carried out. In response to acid stress, cell envelope regulated the membrane fatty acid composition and chain length to reduce damage. Moreover, intracellular pH homeostasis was maintained by increasing H+-ATPase activity and intracellular ATP and amino acid (mainly arginine, glutamate, aspartate and lysine) concentrations. Transcriptional analysis based on RNA-sequencing indicated that acid stress aroused global changes and the differentially expressed genes involved in transcriptional regulation, stress-response protein, transporter, cell envelope, secondary metabolite biosynthesis, DNA and RNA metabolism and ribosome subunit. Consequently, the ATR of S. albulus was preliminarily proposed. Notably, it is indicated that the biosynthesis of ε-PL is also a response mechanism for S. albulus to combat acid stress. These results provide new insights into the ATR of S. albulus and will contribute to the production of ε-PL via adaptive evolution or metabolic engineering.

Identification of the Potential Biological Preservative Tetramycin A-Producing Strain and Enhancing Its Production.

The aim of this study was to develop a potential microbial preservative to prevent the growth of fungi in food. The isolate ZC-G-5 showed strong antifungal activity against food spoilage fungi and Streptomyces albulus was identified on the basis of morphologic, culture, and 16S rDNA sequence analyses. The active metabolite was elucidated as tetramycin A (TMA) through spectroscopic techniques, including HR-ESI-MS, 1D-NMR, and 2D-NMR. An antifungal activity assay revealed that the minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) of TMA were 1.50–2.50 and 3.00–5.00 μg/ml, respectively. In situ antifungal activity analyses demonstrated that 90.0 μg/ml of TMA could inhibit the growth of fungi for over 14 days. In order to enhance TMA production, the high-yield mutant strain YB101 was screened, based on the isolate ZC-G-5, using a high-throughput screening method. The best metabolic precursor was selected during fermentation, when the concentration of glycerol was 8% (v/v) in Gauze’s broth medium to cultivate the mutant strain YB101; the concentration of TMA could be increased to 960.0 μg/ml, compared with the original isolate ZC-G-5, where the concentration of the TMA was only 225.0 μg/ml. Our study may contribute to the application of S. albulus and its active metabolite as a potential bio-preservative in the food industry.

Understanding high ε-poly-l-lysine production by Streptomyces albulus using pH shock strategy in the level of transcriptomics

ε-Poly-L-lysine (ε-PL) is a natural food preservative, which exhibits antimicrobial activity against a wide spectra of microorganisms. The production of ε-PL was significantly enhanced by pH shock in our previous study, but the underlying mechanism is poorly understood. According to transcriptional and physiological analyses in this study, the mprA/B and pepD signal transduction system was first proved to be presented and activated in Streptomyces albulus M-Z18 by pH shock, which positively regulated the transcription of ε-PL synthetase (Pls) gene and enhanced the Pls activity during fermentation. Furthermore, pH shock changed the ratio of unsaturation to saturation fatty acid in the membrane through up-regulating the transcription of fatty acid desaturase genes (SAZ_RS14940, SAZ_RS14945). In addition, pH shock also enhanced the transcription of cytochrome c oxidase (SAZ_RS15070, SAZ_RS15075), ferredoxin reductase (SAZ_RS34975) and iron sulfur protein (SAZ_RS31410) genes, and finally resulted in the improvement of cell respiratory activity. As a result, pH shock was considered to influence a wide range of proteins including regulators, fatty acid desaturase, respiratory chain component, and ATP-binding cassette transporter during fermentation. These combined influences might contribute to enhanced ε-PL productivity with pH shock.

Draft Genome Sequence of the Most Traditional ε-Poly-l-Lysine Producer, Streptomyces albulus NBRC14147.

The genus Streptomyces is known for its secondary metabolite biosynthetic capacities. We report here the draft genome sequence of the most extensively studied ε-poly-l-lysine producer, Streptomyces albulus NBRC14147. Bioinformatic analysis of the 9.6-Mb chromosome identified a large number of secondary metabolite biosynthetic gene clusters.

Discovery of a Short-Chain ε-Poly-l-lysine and Its Highly Efficient Production via Synthetase Swap Strategy.

ε-Poly-l-lysine (ε-PL) is a natural antimicrobial cationic peptide, which is generally recognized as safe for use as a food preservative. To date, the production capacity of strains that produce low-molecular weight ε-PL remains very low and thus unsuitable for industrial production. Here, we report a new low-molecular weight ε-PL-producing Kitasatospora aureofaciens strain. The ε-PL synthase gene of this strain was cloned into a high ε-PL-producing Streptomyces albulus strain. The resulting recombinant strain efficiently produced ε-PL with a molecular weight of 1.3-2.3 kDa and yielded of 23.6 g/L following fed-batch fermentation in a 5 L bioreactor. In addition, circular dichroism spectra showed that this ε-PL takes on a conformation similar to an antiparallel pleated-sheet. Moreover, it demonstrated better antimicrobial activity against yeast compared to the 3.2-4.5 kDa ε-PL. This study provides a highly efficient strategy for production of the low-molecular weight ε-PL, which helps to expand its potential applications.