Biotechnology Letters最新文献

In this study, medlar polyphenol oxidase (PPO) was partially purified by (NH₄)₂SO₄ precipitation and dialysis, respectively. The aim of the study was to investigate the inhibition effects of amino acids, which are candidate PPO ligands, on the activity of the medlar PPO enzyme for advanced biochemical purification techniques and to create a usage field for the enzyme inhibitors in different industrial sectors. No any inhibition studies of amino acids have been investigated on medlar PPO in literature yet. Inactivation of PPO is preferred to be prevention of decreasing of nutritional quality and shelf life of foods. Two bands were determined in electrophoresis analyses. Following, amino acids effects were studied on medlar PPO activity to investigate the potentials of Glycine (Gly), L-Phenylalanine (L-Phe), L-Tyrosine (L-Tyr), L-Cysteine (L-Cys), L-Serine (L-Ser), L-Aspartic acid (L-Asp), L-Histidine (L-His), L-Lysine (L-Lys), L-Proline (L-Pro), and L-Methionine (L-Met) whether acting as natural PPO inhibitors. Inhibition types were determined for catechol and L-Cys was found as a potent competitive inhibitor of medlar PPO. While Gly, L-Phe, L-Pro, L-Ser, L-His, and L-Lys showed uncompetitive inhibition; L-Tyr, L-Asp, and L-Met showed mixed-type inhibition. Statistical analysis was performed to understand whether the chemical structure or concentration of inhibitors showing the same type of inhibition made a statistically significant difference on the enzyme activity %. The results showed that the structure of inhibitors did not make a statistically significant difference on the enzyme activity % while inhibitor concentration created significant difference.

Yeast plays a pivotal role in beer brewing, as its metabolic activity directly determines the flavor profile, product quality, and production efficiency of beer. With the rapid advancement of biotechnology, innovative techniques such as omics, adaptive evolution, and CRISPR-based genome editing have significantly accelerated the process of yeast strain breeding. These technologies not only enhance fermentation performance but also enable the targeted development of novel strains with specific phenotypic traits, thereby addressing diverse market demands for customized beer characteristics. This review systematically discusses current strategies for beer yeast breeding, with particular emphasis on recent technological breakthroughs in strain development. Furthermore, we provide insights into future trends in strain enhancement technologies, highlighting the importance of multidimensional strategies, high-throughput selection platforms, the synergistic integration of synthetic biology and computational modeling to achieve precise strain optimization. This review highlights that continuous technological innovation is crucial for enhancing yeast breeding efficiency and meeting the evolving demands of the industry.

D-Asp is a pharmaceutically valuable amino acid used as an antibiotic precursor and has potential therapeutic benefits for neurological disorders and reproductive health. The lactic acid bacterium Latilactobacillus curvatus strain WDN19, originally isolated from a salt-fermented pickle, produces large amounts of D-Asp from L-Asp. To optimize D-Asp production, we investigated the effects of various cultivation conditions. Growth and D-Asp formation increased concomitantly with temperature, reaching a maximum at 37 °C, whereas no growth occurred at 40 °C. L-Asp concentrations ≤ 100 mM minimally affected biomass, yet higher levels curtailed both growth and conversion, revealing an intrinsic racemization capacity of ~ 30 mM under unbuffered conditions. Strain WDN19 grew across an initial pH range of 5.0-9.5, but rapid acidification to pH ≈ 4.5 triggered severe cell death and arrested D-Asp synthesis. Incorporation of 500 mM 2-morpholinoethanesulfonic acid maintained near-neutral pH, preserved viability, and enabled almost quantitative racemization of up to 500 mM L-Asp within 48 h, yielding > 250 mM D-Asp. These findings establish pH control as the critical determinant for high-yield D-Asp production by strain WDN19, providing a practical framework for environmentally friendly, fermentation-based manufacturing of this pharmaceutically valuable amino acid.

This study focused on phytase overproduction in recombinant E. coli BL21(DE3)/pET-appA expressing E. coli phytase(APPA). A modified mineral salt medium was investigated for the strain growth and phytase production in shake-flask, which included 30 mM NH₄OH and 17 g L^-1 KH₂PO₄ as sole nitrogen(N) and phosphorous(P) sources, respectively. After that, fed-batch cultivation process, especially induction strategy feeding lactose intermittently was optimized in 30L bioreactor by Taguchi orthogonal experiments (L₁₈(2¹ × 3⁶)) at an induction cell density of OD₆₀₀ 25, followed by research on effect of induction cell densities. The optimized fed-batch cultivation process parameters included glucose starvation time of 15 min just before induction, induction cell density of OD₆₀₀ 45, induction temperature of 25 °C, predetermined specific growth rate of 0.04 h^-1, DO level of 15%, lactose concentration of 60 mM, lactose feed times of 6 times, and pH of 7.2-7.6. In the optimized condition, the highest soluble phytase activity of 2 152U mL^-1 ever reported was obtained at 33.0 h, indicating the enhancements of 31.6-fold and 3.7-fold compared to batch cultivation process in shake flask and pre-optimized fed-batch cultivation process in the bioreactor, respectively. We propose that the defined medium, the induction strategy and fed-batch process parameters could be applied to overproduce soluble proteins in E. coli.

The global inclination in the production of building block chemicals is shifting in the direction of sourcing sustainable resources and implementing circular economy frameworks. Biotechnological approaches facilitate the economic advancement of precursor production through the valorization of waste biomass while alleviating the impacts on ecology. Strategies in white biotechnology enable the utilization of affordable and renewable resources for the synthesis of a wide spectrum of bio-based compounds. Considering the increasing market demand for 2-keto-D-gluconic acid and its derivatives, this review provides a systematic overview of its physicochemical properties and applications across different industries. This paper critically compares chemical and microbial production methods, with a particular focus on the microorganisms employed, the substrates utilized and the mode of operation. Notably, this review covers emerging trends in the utilization of agro-industrial residues as substrates and highlights advances in microbial strain engineering and process optimization. In addition to providing a comprehensive overview of the critical aspects of 2-KGA production, this article underscores the key challenges and outlines necessary advancements. This study intends to steer future research toward optimizing 2-KGA production processes for more environmentally sustainable methods.

Astaxanthin is a valuable carotenoid with potent antioxidant properties and has broad applications in the pharmaceutical, nutraceutical, and cosmetic industries. In this study, Rhodotorula toruloides CB6-10/1, isolated from Canna indica L. flowers, was evaluated for astaxanthin production. The orange-red pigment was confirmed as astaxanthin via thin-layer chromatography and high-performance liquid chromatography, with quantification performed by spectrophotometry. Comprehensive genome analysis and production optimization of R. toruloides CB6-10/1 confirmed the presence of key astaxanthin biosynthesis genes, such as CrtYB, CrtI, CrtW, CrtZ, and CrtR, which facilitate the conversion of β-carotene to astaxanthin through hydroxylation and ketolation. The key parameters, including carbon and nitrogen sources, their concentrations, trace elements, agitation speed, and pH, were systematically evaluated to optimize production. Although copper appeared beneficial in the Plackett-Burman screening, its effect and those of other metals were not significant. Optimization using Response Surface Methodology for cost-effective nitrogen sources determined that a combination of 1.10 g/L yeast extract, 10.0 g/L peptone, and 0.50 g/L ammonium sulfate yielded the maximum astaxanthin production of 4.728 mg/L, under cultivation conditions of pH 4.5, 200 rpm, and 30 g/L glucose, representing a fourfold increase compared with the basal medium. This optimization not only enhances pigment production efficiency but also reduces dependency on costly trace elements, improving process scalability and economic feasibility. Overall, these results demonstrate R. toruloides CB6-10/1 as a promising microbial source for sustainable astaxanthin production with potential further applications.

Cellulase-driven enzymatic hydrolysis of lignocellulosic biomass plays a crucial role in its bioconversion into renewable biofuels. The present study aimed to identify soil-borne microfungal isolates for their cellulase producing ability. Total of eleven species were selected based on their hydrolytic screening assay. The selected isolates were cultivated on four different substrates supplemented czapeck broth (i.e., corncob, wheat straw, black poplar wood, and CMC) under varying conditions of temperature, incubation time, and pH. The data were analyzed using two-factor analysis of variance (ANOVA) with interaction effect. The results revealed highly significant variability in enzyme activities in response to different substrates, pH levels, temperatures, and incubation periods. Among the tested isolates, Fusarium sp. (LMPP-24-S2) exhibited the highest endoglucanase (1.84 U/mL) and exoglucanase (2.17 U/mL) activities, while Penicellium oxalicum showed the highest β-glucosidase activity (9.09 U/mL) in corncob medium. These findings suggest that Fusarium sp. (LMPP-24-S2) and P. oxalicum are efficient producers of cellulolytic enzymes and that corncob serves as an effective substrate for an optimal cellulase induction. Maximum productivity was achieved at 30 °C, pH 5, and a 7-day incubation period. Especially, four isolates of this study comprising Clonostachys epichloë, Stagonosporopsis cucurbitacearum, Bipolaris cynodontis, and Pyrenochaetopsis leptospora were reported for the first time for their cellulase-producing potential.

A DNA marker is routinely used to determine the size of DNA fragments by electrophoresis in molecular biology laboratories. Thus, in-house produced DNA markers could be appropriate for moderately equipped laboratories. We report here a new procedure for customizing fragment sizes of DNA markers and then generating 9 fragments of DNA markers, ranging from 500 to 4600 bp, by partial digestion of the recombinant pSY4.6-1 and pSY4.6-2 plasmids using only one restriction enzyme. DNA markers, containing 9 fragments, were obtained by digesting the recombinant plasmid, which resulted from the commercial cloning pJET1.1 and pJET1.2 plasmids and inserts of 1506 bp or 1628 bp, and one suitable restriction enzyme. Our in-house produced DNA markers, containing desirable fragment sizes, could be suitable for moderately equipped laboratories due to their simplicity, time-saving, and cost-saving procedure (approximately US$5 for materials to produce a DNA marker used for 100 agarose gel lanes) compared to the current ones widely used in most laboratories.

The widespread use and environmental persistence of polyethylene (PE) have led to a global pollution crisis, which is intensified by its fragmentation into hazardous micro- and nanoplastics. Although bioremediation using polymer-degrading microbes presents a sustainable alternative, only a limited microbes have been identified, primarily due to the challenges of culturing potential degraders in the laboratory. We isolated two P. aeruginosa strains (SKD-SD-3171 and SS-SD-7780) from urban waste disposal areas and comparatively evaluated their PE degrading efficacy, over 120 days. Both strains utilized PE as a carbon source, as confirmed by weight loss (24.53±0.35% for P. aeruginosa SKD-SD-3171 and 22.50±0.35% for P. aeruginosa SS-SD-7780), polymer reduction rate (K=0.00235 day^-1±0.00004 for P. aeruginosa SKD-SD-3171 and 0.00212 day^-1±0.00004 for P. aeruginosa SS-SD-7780), and calculation of half-life (t1/2=295.55±4.77 for P. aeruginosa SKD-SD-3171 and 326.39±5.94 for P. aeruginosa SS-SD-7780) after the incubation in a carbon-free medium. Biodegradation was further validated using fourier transform infrared spectroscopy (FT-IR), which showed structural alterations, and field emission scanning electron microscopy (FE-SEM), which revealed surface erosion in PE, following microbial treatment. Additionally, gas chromatography-mass spectrometry (GC-MS) identified degradation intermediates whose kinetic profiling revealed effective polyethylene degradation through biodegradation efficiency metrics. Our findings demonstrate that P. aeruginosa SKD-SD-3171 exhibits comparatively faster and consistent polyethylene degradation kinetics than P. aeruginosa SS-SD-7780 under laboratory-based conditions and it also establishes effective methodological framework for isolation, selection and evaluation of specific polymer-degrading microorganisms using advanced analytical techniques. These findings provide insights into developing PE-waste management strategies like enzyme characterization, followed by field-scale validation to enhance degradation kinetics.