Microbial Cell Factories最新文献

This comprehensive review explores the emergence of titanium dioxide nanoparticles (TiO₂-NPs) as versatile nanomaterials, particularly exploring their biogenic synthesis methods through different biological entities such as plants, bacteria, fungi, viruses, and algae. These biological entities provide eco-friendly, cost-effective, biocompatible, and rapid methods for TiO₂-NP synthesis to overcome the disadvantages of traditional approaches. TiO₂-NPs have distinctive properties, including high surface area, stability, UV protection, and photocatalytic activity, which enable diverse applications. Through detailed analysis, this review demonstrates significant applications of green fabricated TiO₂-NPs in biomedicine, explicitly highlighting their antimicrobial, anticancer, and antioxidant activities, along with applications in targeted drug delivery, photodynamic therapy, and theragnostic cancer treatment. Additionally, the review underscores their pivotal significance in biosensors, bioimaging, and agricultural applications such as nanopesticides and nanofertilizers. Also, this review proves valuable incorporation of TiO₂-NPs in the treatment of contaminated soil and water with various environmental contaminants such as dyes, heavy metals, radionuclides, agricultural effluents, and pathogens. These comprehensive findings establish the foundation for future innovations in nanotechnology, underscoring the importance of further investigating bio-based synthetic approaches and bioactivity mechanisms to enhance their efficacy and safety across healthcare, agricultural, and environmental applications.

Bacterial biofilms pose significant challenges, from healthcare-associated infections to biofouling in industrial systems, resulting in significant health impacts and financial losses globally. Classic antimicrobial methods often fail to eradicate sessile microbial communities within biofilms, requiring innovative approaches. This review explores the structure, formation, and role of biofilms, highlighting the critical importance of exopolysaccharides in biofilm stability and resistance mechanisms. We emphasize the potential of microbial enzymatic approaches, particularly focusing on glycosidases, proteases, and deoxyribonucleases, which can disrupt biofilm matrices effectively. We also delve into the importance of enzymes such as cellobiose dehydrogenase, which disrupts biofilms by degrading polysaccharides. This enzyme is mainly sourced from Aspergillus niger and Sclerotium rolfsii, with optimized production strategies enhancing its efficacy. Additionally, we explore levan hydrolase, alginate lyase, α-amylase, protease, and lysostaphin as potent antibiofilm agents, discussing their microbial origins and production optimization strategies. These enzymes offer promising avenues for combating biofilm-related challenges in healthcare, environmental, and industrial settings. Ultimately, enzymatic strategies present environmentally friendly solutions with high potential for biofilm management and infection control.

The conversion of CO₂ into methanol depicts one of the most promising emerging renewable routes for the chemical and biotech industry. Under this regard, native methylotrophs have a large potential for converting methanol into value-added products but require targeted engineering approaches to enhance their performances and to widen their product spectrum. Here we use a systems-based approach to analyze and engineer M. extorquens TK 0001 for production of glycolic acid. Application of constraint-based metabolic modeling reveals the great potential of M. extorquens for that purpose, which is not yet described in literature. In particular, a superior theoretical product yield of 1.0 C-mol_{Glycolic acid} C-mol_Methanol^-1 is predicted by our model, surpassing theoretical yields of sugar fermentation. Following this approach, we show here that strain engineering is viable and present 1st generation strains producing glycolic acid via a heterologous NADPH-dependent glyoxylate reductase. It was found that lactic acid is a surprising by-product of glycolic acid formation in M. extorquens, most likely due to a surplus of available NADH upon glycolic acid synthesis. Finally, the best performing strain was tested in a fed-batch fermentation producing a mixture of up to total 1.2 g L^-1 glycolic acid and lactic acid. Several key performance indicators of our glycolic acid producer strain are superior to state-of-the-art synthetic methylotrophs. The presented results open the door for further strain engineering of the native methylotroph M. extorquens and pave the way to produce two promising biopolymer building blocks from green methanol, i.e., glycolic acid and lactic acid.

Background: Universal stress proteins (USPs) are prevalent in various bacteria to cope with different adverse stresses, while their possible effects on secondary metabolisms of hosts are unclear. Tiancimycins (TNMs) are ten-membered endiynes possessing excellent potential for development of anticancer antibody-drug conjugates. During our efforts to improve TNMs titer, a high-producing strain Streptomyces sp. CB03234-S had been obtained and its possible high yield mechanism is being continuously explored to further enhance TNMs production.

Results: In this work, the whole-genome resequencing and analysis results revealed a notable 583 kb terminal deletion containing 8 highly expressed usp genes in the genome of CB03234-S. The individual complementation of lost USPs in CB03234-S all showed differential effects on secondary metabolism, especially TNMs production. Among them, the overexpression of USP3 increased TNMs titer from 12.8 ± 0.2 to 31.1 ± 2.3 mg/L, while the overexpression of USP8 significantly reduced TNMs titer to only 1.0 ± 0.1 mg/L, but activated the production of porphyrin-type compounds. Subsequent genetic manipulations on USP3/USP8 orthologs in Streptomyces. coelicolor A3(2) and Streptomyces sp. CB00271 also presented clear effects on the secondary metabolisms of hosts. Further sequence similarity network analysis and Streptomyces-based pan‑genomic analysis suggested that the USP3/USP8 orthologs are widely distributed across Streptomyces.

Conclusion: Our studies shed light on the potential effects of USPs on secondary metabolisms of streptomycetes for the first time, and USPs could become novel targets for exploring and exploiting natural products in streptomycetes.

Electrospun nanofibers offer a highly promising platform for the delivery of vaginal lactobacilli, providing an innovative approach to preventing and treating vaginal infections. To advance the application of nanofibers for the delivery of lactobacilli, tools for studying their safety and efficacy in vitro need to be established. In this study, fluorescent (mCherry and GFP) and luminescent (NanoLuc luciferase) proteins were expressed in three vaginal lactobacilli (Lactobacillus crispatus, Lactobacillus gasseri and Lactobacillus jensenii) and a control Lactiplantibacillus plantarum with the aim to use this technology for close tracking of lactobacilli release from nanofibers and their adhesion on epithelial cells. The recombinant proteins influenced the growth of the bacteria, but not their ability to produce hydrogen peroxide. Survival of lactobacilli in nanofibers immediately after electrospinning varied among species. Bacteria retained fluorescence upon incorporation into PEO nanofibers, which was vital for evaluation of their rapid release. In addition, fluorescent labelling facilitated efficient tracking of bacterial adhesion to Caco-2 epithelial cells, while luminescence provided important quantitative insights into bacterial attachment, which varied from 0.5 to 50% depending on the species. The four lactobacilli in dispersion or in nanofibers were not detrimental for the viability of Caco-2 cells, and did not demonstrate hemolytic activity highlighting the safety profiles of both bacteria and PEO nanofibers. To summarize, this study contributes to the development of a promising delivery system, tailored for local administration of safe vaginal lactobacilli.

Background: Biotechnologies that utilize microorganisms as production hosts for lipid synthesis will enable an efficient and sustainable solution to produce lipids, decreasing reliance on traditional routes for production (either petrochemical or plant-derived) and supporting a circular bioeconomy. To realize this goal, continuous biomanufacturing processes must be developed to maximize productivity and minimize costs compared to traditional batch fermentation processes.

Results: Here, we utilized biofilms of the marine bacterium, Marinobacter atlanticus, to produce wax esters from succinate (i.e., a non-sugar feedstock) to determine its potential to serve as a production chassis in a continuous flow, biofilm-based biomanufacturing process. To accomplish this, we evaluated growth as a function of protein concentration and wax ester production from M. atlanticus biofilms in a continuously operated 3-D printed fixed bed bioreactor. We determined that exposing M. atlanticus biofilms to alternating nitrogen-rich (1.8 mM NH₄⁺) and nitrogen-poor (0 mM NH₄⁺) conditions in the bioreactor resulted in wax ester production (26 ± 5 mg/L, normalized to reactor volume) at a similar concentration to what is observed from planktonic M. atlanticus cells grown in shake flasks previously in our lab (ca. 25 mg/L cell culture). The wax ester profile was predominated by multiple compounds with 32 carbon chain length (C₃₂; 50-60% of the total). Biomass production in the reactor was positively correlated with dilution rate, as indicated by protein concentration (maximum of 1380 ± 110 mg/L at 0.4 min^-1 dilution rate) and oxygen uptake rate (maximum of 4 mmol O₂/L/h at 0.4 min^-1 dilution rate) measurements at different flow rates. Further, we determined the baseline succinate consumption rate for M. atlanticus biofilms to be 0.16 ± 0.03 mmol/L/h, which indicated that oxygen is the limiting reactant in the process.

Conclusion: The results presented here are the first step toward demonstrating that M. atlanticus biofilms can be used as the basis for development of a continuous flow wax ester biomanufacturing process from non-sugar feedstocks, which will further enable sustainable lipid production in a future circular bioeconomy.

Microbial exopolysaccharides (EPSs) possess valuable biological functions and fascinating physicochemical properties. On the other hand, lung cancer is the primary contributor to global cancer-related deaths. However, health and safety concerns have prevented the identification and approval of any medications, including chemotherapeutic agents, for lung cancer treatment to date. The current study aims to enhance the production of bacterial EPS as a coating agent for the synthesis of selenium nanoparticles (AZEPS-SeNPs), to enhance their biological activity against pathogenic microbes, human lung adenocarcinoma cells (A549) in vitro, and diethyl nitrosamine (DEN)-induced lung cancer in vivo. The synthesized AZEPS-SeNPs exhibited a significant antifungal effect reaching 49.3 mm against Candida albicans. SeNPs and EPSs demonstrated a concentration-dependent synergistic antioxidant effect of 96.8%. Moreover, the synthesized nanoparticles showed a highly potent cytotoxic effect against A549 cells (1.724 ± 0.08 µg/mL) with a therapeutic index of 7.18 ± 0.21 that leads to increased reactive oxygen species (ROS) production. AZEPS-SeNPs demonstrated a proapoptotic effect on the lung adenocarcinoma A549 cell line by stimulating caspase 3 and Bax (7.08-fold and 6.505-fold, respectively), inhibiting the anti-apoptotic gene Bcl2, and arresting the cell cycle in the S phase. In vivo study revealed that the AZEPS-SeNPs-treated group showed improved histopathological examination of lung tissue sections. The present study concluded the efficiency of the synthesized bacterial EPS-SeNPs as multi-functional antimicrobial, anticancer and antioxidant agent.

Background: Bacillus subtilis is widely used for industrial enzyme production due to its capacity to efficiently secrete proteins. However, secretion efficiency of enzymes varies widely, and optimizing secretion is crucial to make production commercially viable. Previously, we have shown that overexpression of the xylanase XynA lowers expression of Clp protein chaperones, and that inactivation of CtsR, which regulates and represses clp transcription, increases the production of XynA. In the current study, we examined whether the same is the case for overexpression of the α-amylase AmyM from Geobacillus stearothermophilus by B. subtilis, and why XynA shows a different timing of secretion compared to AmyM.

Results: Transcriptome analyses revealed that B. subtilis cells overexpressing AmyM exhibited a distinct profile compared to XynA overexpressing cells, however there were also similarities and in both cases expression of CtsR controlled genes was downregulated. In contrast to XynA, inactivation of CtsR did not improve AmyM production. Upregulation of other protein chaperones, including GroEL/ES and DnaJ/K, by inactivating their transcriptional repressor HrcA, had almost no effect on XynA yields and in fact considerably lowered that of AmyM. Despite using the same promoter, the production of XynA peaks well before AmyM reaches its optimal secretion rate. Transcriptome and ribosome profiling indicated that this is neither related to transcription nor to translation regulation. We show that the reduced secretion in the stationary phase is partially due to the activity of secreted proteases, but also due to the activity of the intracellular protease LonA. The absence of this protein resulted in a 140% and 20% increased production for XynA and AmyM, respectively.

Conclusion: The combination of transcriptome and ribosome profiling offered important information to determine at which cellular level production bottlenecks occurred. This helped us to identify LonA protease as an important factor influencing enzyme production yields in B. subtilis.

Fungi can synthesize a diverse range of melanins with appropriate physicochemical and biological characteristics for numerous applications in health, environmental protection, energy, and industry. Gaining deeper insights into the chemical structures, biosynthetic pathways, and regulatory mechanisms of fungal melanin would establish a basis for metabolic engineering approaches, aimed at enhancing production efficiency and creating custom-designed melanin with desirable material properties. Due to growing interest in their beneficial effects and applications, research on the structure, biosynthesis, and regulation of fungal melanin has significantly advanced. This review highlighted recent progress in fungal melanin production and applications, concentrating on structure, biosynthesis, and regulatory networks, and suggested how an improved understanding of melanin biosynthesis could enable efficient production for future applications.

L-Homoserine, serves as a non-essential precursor for the essential amino acids derived from L-aspartate in both animals and humans. It finds widespread applications across the food, cosmetics, pharmaceutical, and animal feed industries. Microbial fermentation, primarily utilizing Escherichia coli, is the dominant approach for L-Homoserine production. However, despite recent advancements in fermentative processes employing E. coli strains, low production efficiency remains a significant barrier to its commercial viability. This review explores the biosynthesis, secretion, and regulatory mechanisms of L-Homoserine in E. coli while assessing various metabolic engineering strategies aimed at improving production efficiency.