Establishment of LAMP-CRISPR/Cas12a for rapid detection of Escherichia coli O157:H7 and one-pot detection.

Escherichia coli O157:H7 is a pathogenic serotype of Escherichia coli. Consumption of food contaminated with E. coli O157:H7 could cause a range of diseases. Therefore, it is of great importance to establish rapid and accurate detection methods for E. coli O157:H7 in food. In this study, based on LAMP and combined with the CRISPR/cas12a system, a sensitive and specific rapid detection method for E. coli O157:H7 was established, and One-Pot detection method was also constructed. The sensitivity of this method could stably reach 9.2 × 10° CFU/mL in pure culture, and the whole reaction can be completed within 1 h. In milk, E. coli O157:H7 with an initial contamination of 7.4 × 10° CFU/mL only needed to be cultured for 3 h to be detected. The test results can be judged by the fluorescence curve or by visual observation under a UV lamp, eliminating instrument limitations and One-Pot detection can effectively prevent the problem of false positives. In a word, the LAMP-CRISPR/cas12a system is a highly sensitive and convenient method for detecting E. coli O157:H7.

Fermentation of millet bran with Bacillus natto: enhancement of bioactivity levels and the bioactivity of bran extract.

BACKGROUND: Millet bran (MB), a byproduct of millet production, is rich in functional components but it is underutilized. In recent years, researchers have shown that fermentation can improve the biological activity of cereals and their byproducts. This study used Bacillus natto to ferment millet bran to improve its added value and broaden the application of MB. The bioactive component content, physicochemical properties, and functional activity of millet bran extract (MBE) from fermented millet bran were determined. RESULTS: After fermentation, the soluble dietary fiber (SDF) content increased by 92.0%, the ß-glucan content by 164.4%, the polypeptide content by 111.4%, the polyphenol content by 32.5%, the flavone content by 16.4%, and the total amino acid content by 95.4%. Scanning electron microscopy revealed that the microscopic morphology of MBE changed from complete and dense blocks to loosely porous shapes after fermentation. After fermentation, the solubility, water-holding capacity, and viscosity significantly increased and the particle size decreased. Moreover, the glucose adsorption capacity (2.1 mmol g-1), glucose dialysis retardation index (75.3%), and α-glucosidase inhibitory (71.4%, mixed reversible inhibition) activity of the fermented MBE (FMBE) were greater than those of the unfermented MBE (0.99 mmol g-1, 32.1%, and 35.1%, respectively). The FMBE presented better cholesterol and sodium cholate (SC) adsorption properties and the adsorption was considered inhomogeneous surface adsorption. CONCLUSION: Fermentation increased the bioactive component content and improved the physicochemical properties of MBE, thereby improving its hypoglycemic and hypolipidemic properties. This study not only resolves the problem of millet bran waste but also encourages the development of higher value-added application methods for millet bran. © 2024 Society of Chemical Industry.

Effects of monolauroyl-galactosylglycerol on membrane fatty acids and properties of Bacillus cereus.

The purpose of this study was to provide new ideas for the antibacterial mechanism of monolauroyl-galactosylglycerol (MLGG) from the perspective of cell membranes. The changes in cell membrane properties of Bacillus cereus (B. cereus) CMCC 66,301 exposed to different concentrations (1 × MIC (minimum inhibitory concentration), 2 × MIC, 1 × MBC (minimum bacterial concentration)) of MLGG were evaluated. It was found that the lag phase of B. cereus cells was prolonged at low concentration MLGG (1 × MIC and 2 × MIC), while about 2 log CFU/mL reduction in B. cereus populations were observed when exposed to high concentration MLGG (1 × MBC). MLGG treated B. cereus displayed obvious membrane depolarization, while membrane permeability had no change using PI (propidium iodide) staining. Significant increase in the membrane fluidity in response to MLGG exposure occurred, which was consistent with the modification of membrane fatty acids compositions, where the relative content of straight-chain fatty acids (SCFAs) and unsaturated fatty acids (UFAs) increased, while branched-chain fatty acids (BCFAs) decreased significantly. The decreased transition Tm value and cell surface hydrophobicity was also observed. Additionally, effect of MLGG on bacterial membrane compositions were explored at the submolecular level by infrared spectroscopy. Resistance tests of B. cereus to MLGG had demonstrated the advantages of MLGG as a bacteriostatic agent. Collectively, these studies indicate that modifying the fatty acid composition and properties of cellular membranes through MLGG exposure is crucial for inhibiting bacteria growth, providing new insights into the antimicrobial mechanisms of MLGG. KEY POINTS: • Monolauroyl-galactosylglycerol inserted into B. cereus lipid bilayer membrane • Monolauroyl-galactosylglycerol treatment caused B. cereus membrane depolarization • Monolauroyl-galactosylglycerol resulted in B. cereus membrane fatty acids alteration.

Acetate and auto-inducing peptide are independent triggers of quorum sensing in Lactobacillus plantarum.

The synthesis of plantaricin in Lactobacillus plantarum is regulated by quorum sensing. However, the nature of the extra-cytoplasmic (EC) sensing domain of the histidine kinase (PlnB1) and the ability to recognize the auto-inducing peptide PlnA1 is not known. We demonstrate the key motif Ile-Ser-Met-Leu of auto-inducing peptide PlnA1 binds to the hydrophobic region Phe-Ala-Ser-Gln-Phe of EC loop 2 of PlnB1 via hydrophobic interactions and hydrogen bonding. Moreover, we identify a new inducer, acetate, that regulates the synthesis of plantaricin by binding to a positively charged region (Arg-Arg-Tyr-Ser-His-Lys) in loop 4 of PlnB1 via electrostatic interaction. The side chain of Phe143 on loop 4 determined the specificity and affinity of PlnB1 to recognize acetate. PlnA1 activates quorum sensing in log phase growth and acetate in stationary phase to maintain the synthesis of plantaricin under conditions of reduced growth. Acetate activation of PlnB was also evident in four types of PlnB present in different Lb. plantarum strains. Finally, we proposed a model to explain the developmental regulation of plantaricin synthesis by PlnA and acetate. These results have potential applications in improving food fermentation and bacteriocin production.

Shared signaling pathways and targeted therapy by natural bioactive compounds for obesity and type 2 diabetes.

Epidemiological evidence showed that patients suffering from obesity and T2DM are significantly at higher risk for chronic low-grade inflammation, oxidative stress, nonalcoholic fatty liver (NAFLD) and intestinal flora imbalance. Increasing evidence of pathological characteristics illustrates that some common signaling pathways participate in the occurrence, progression, treatment, and prevention of obesity and T2DM. These signaling pathways contain the pivotal players in glucose and lipid metabolism, e.g., AMPK, PI3K/AKT, FGF21, Hedgehog, Notch, and WNT; the inflammation response, for instance, Nrf2, MAPK, NF- kB, and JAK/STAT. Bioactive compounds from plants have emerged as key food components related to healthy status and disease prevention. They can act as signaling molecules to initiate or mediate signaling transduction that regulates cell function and homeostasis to repair and re-functionalize the damaged tissues and organs. Therefore, it is crucial to continuously investigate bioactive compounds as sources of new pharmaceuticals for obesity and T2DM. This review provides comprehensive information of the commonly shared signaling pathways between obesity and T2DM, and we also summarize the therapeutic bioactive compounds that may serve as anti-obesity and/or anti-diabetes therapeutics by regulating these associated pathways, which contribute to improving glucose and lipid metabolism, attenuating inflammation.

An endolysin Salmcide-p1 from bacteriophage fmb-p1 against gram-negative bacteria.

AIMS: A novel endolysin Salmcide-p1 was developed as a promising candidate of new preservative and a supplement to effective enzyme preparations against gram-negative bacterial contaminations. METHODS AND RESULTS: Salmcide-p1 was identified by complementing the genomic sequence of a virulent Salmonella phage fmb-p1. Salmcide-p1 of 112 µg ml-1 could quickly kill Salmonella incubated with 100 mmol l-1 EDTA, with no haemolytic activity. Meanwhile, Salmcide-p1 had a high activity of lysing Salmonella cell wall peptidoglycan. At different temperatures (4-75°C), pH (4-11) and NaCl concentration (10-200 mmol l-1 ), the relative activity of Salmcide-p1 was above 60%. At 4°C, the combination of Salmcide-p1 and EDTA-2Na could inhibit the number of Salmonella Typhimurium CMCC 50115 in skim milk to less than 4 log CFU ml-1 by 3 days, and the number of Shigella flexneri CMCC 51571 was lower than 4 log CFU ml-1 by 9 days. CONCLUSIONS: Salmcide-p1 had a wide bactericidal activity against gram-negative bacteria and showed a broader anti-Salmonella spectrum than the phage fmb-p1. The combination strategy of Salmcide-p1 and EDTA-2Na could significantly inhibit the growth of gram-negative bacteria inoculated in skim milk. SIGNIFICANCE AND IMPACT OF THE STUDY: Bacteriophage endolysin as an antibacterial agent is considered to be a new strategy against bacterial contamination.

Thermostability Improvement of L-Asparaginase from Acinetobacter soli via Consensus-Designed Cysteine Residue Substitution.

To extend the application range of L-asparaginase in food pre-processing, the thermostability improvement of the enzyme is essential. Herein, two non-conserved cysteine residues with easily oxidized free sulfhydryl groups, Cys8 and Cys283, of Acinetobacter soli L-asparaginase (AsA) were screened out via consensus design. After saturation mutagenesis and combinatorial mutation, the mutant C8Y/C283Q with highly improved thermostability was obtained with a half-life of 361.6 min at 40 °C, an over 34-fold increase compared with that of the wild-type. Its melting temperature (Tm) value reaches 62.3 °C, which is 7.1 °C higher than that of the wild-type. Molecular dynamics simulation and structure analysis revealed the formation of new hydrogen bonds of Gln283 and the aromatic interaction of Tyr8 formed with adjacent residues, resulting in enhanced thermostability. The improvement in the thermostability of L-asparaginase could efficiently enhance its effect on acrylamide inhibition; the contents of acrylamide in potato chips were efficiently reduced by 86.50% after a mutant C8Y/C283Q treatment, which was significantly higher than the 59.05% reduction after the AsA wild-type treatment. In addition, the investigation of the mechanism behind the enhanced thermostability of AsA could further direct the modification of L-asparaginases for expanding their clinical and industrial applications.

Acetate Activates Lactobacillus Bacteriocin Synthesis by Controlling Quorum Sensing.

Bacteriocins are useful for controlling the composition of microorganisms in fermented food. Bacteriocin synthesis is regulated by quorum sensing mediated by autoinducing peptides. In addition, short-chain fatty acids, especially acetic acid, reportedly regulate bacteriocin synthesis. Five histidine kinases that regulated the synthesis of bacteriocins were selected to verify their interactions with acetate. Acetate activated the kinase activity of PlnB, SppK, and HpK3 in vitro and increased the yield of their cognate bacteriocins plantaricin EF, sakacin A, and rhamnosin B in vivo. The antimicrobial activity against Staphylococcus aureus of the fermentation supernatants of Lactobacillus plantarum, Lactobacillus sakei, and Lactobacillus rhamnosus with addition of acetate increased to 298%, 198%, and 289%, respectively, compared with that in the absence of acetate. Our study elucidated the activation activity of acetate in bacteriocin synthesis, and it might provide a potential strategy to increase the production of bacteriocin produced by Lactobacillus. IMPORTANCE Bacteriocins produced by lactic acid bacteria (LAB) are particularly useful in food preservation and food safety. Bacteriocins might increase bacterial competitive advantage against the indigenous microbiota of the intestines; at the same time, bacteriocins could limit the growth of undesired microorganisms in yogurt and other dairy products. This study confirmed that three kinds of histidine kinases were activated by acetate and upregulated bacteriocin synthesis both in vitro and in vivo. The increasing yield of bacteriocins reduced the number of pathogens and increased the number of probiotics in milk. Bacteriocin synthesis activation by acetate may have a broad application in the preservation of dairy products and forage silage.

L. johnsonii, L. plantarum, and L. rhamnosus alleviated Enterohaemorrhagic Escherichia coli-induced diarrhoea in mice by regulating gut microbiota.

Enterohaemorrhagic Escherichia coli (EHEC) is a prominent foodborne pathogen that causes infectious intestinal diarrhoea. Lactobacillus is a recognized probiotic that inhibits intestinal pathogens and maintains the balance of the intestinal flora. The purpose of this study was to investigate the regulatory effects of three Lactobacillus strains, L. johnsonii, L. plantarum, and L. rhamnosus, on the intestinal flora of EHEC-infected mice. The initial weight and diarrhoea index of the mice were recorded. After 21 days, the faeces of the mice were subjected to 16S rDNA high-throughput sequencing. The diarrhoea index of mice treated with Lactobacillus improved, their body weight continued to rise, and their liver index gradually decreased. The α diversity analysis showed that the intestinal flora diversity and abundance were lower in mice infected with EHEC than in healthy mice. L. plantarum, L. johnsonii, and L. rhamnosus significantly improved the diversity of the flora species. In terms of flora composition, the three main phyla present were Bacteroidetes, Firmicutes, and Proteobacteria. The abundance of these three phyla was reduced to 93.81% after infection and restored to over 96.30% after treatment. At the genus level, Lactobacillus reduced the abundance of Bacteroides, Helicobacter pylori, and Shigella, while increasing the abundance of butyric acid-producing bacteria and Lactobacillus. Finally, a heat map and non-metric multidimensional scaling analysis showed that the intestinal flora structures in the L. johnsonii, L. plantarum, and L. rhamnosus treatment groups were closest to those of healthy mice. In conclusion, L. johnsonii, L. plantarum, and L. rhamnosus regulated and improved the structure of intestinal flora and relieved diarrhoea caused by EHEC infection.

Transcriptomic and proteomic profiling response of methicillin-resistant Staphylococcus aureus (MRSA) to a novel bacteriocin, plantaricin GZ1-27 and its inhibition of biofilm formation.

Methicillin-resistant Staphylococcus aureus (MRSA) has become a worrisome superbug, due to its wide distribution and multidrug resistance. To characterize effects of a newly identified plantaricin GZ1-27 on MRSA, transcriptomic and proteomic profiling of MRSA strain ATCC43300 was performed in response to sub-MIC (16 µg/mL) plantaricin GZ1-27 stress. In total, 1090 differentially expressed genes (padj < 0.05) and 418 differentially expressed proteins (fold change > 1.2, p < 0.05) were identified. Centralized protein expression clusters were predicted in biological functions (biofilm formation, DNA replication and repair, and heat-shock) and metabolic pathways (purine metabolism, amino acid metabolism, and biosynthesis of secondary metabolites). Moreover, a capacity of inhibition MRSA biofilm formation and killing biofilm cells were verified using crystal violet staining, scanning electron microscopy, and confocal laser-scanning microscopy. These findings yielded comprehensive new data regarding responses induced by plantaricin and could inform evidence-based methods to mitigate MRSA biofilm formation.

Surfactin effectively inhibits Staphylococcus aureus adhesion and biofilm formation on surfaces.

Biosurfactants are amphiphilic compounds that composed of hydrophilic and hydrophobic moieties, which possess the ability of self-organizing between phases, reducing the interfacial tension, and forming aggregates such as micelles. This spontaneous process results in significant changes in surface properties that directly influence the adherence of microorganisms. In this study, the ability of surfactin, a biosurfactant produced by Bacillus subtilis in reducing adhesion and disrupting the presence of biofilm of Staphylococcus aureus (S. aureus) on several surfaces, was investigated. Significant biofilm removal was observed on glass, polystyrene, and stainless steel surfaces. Furthermore, we explored the probable mechanism about how surfactin affected S. aureus biofilm formation. Based on our findings, surfactin had a significant effect on the polysaccharides production and especially decreased the percentage of alkali-soluble polysaccharide in biofilms. It also down-regulated the expression of icaA and icaD significantly, which are necessary for the important constituents to take shape of staphylococcal biofilm. In addition, it was found that the lipopeptide affected the quorum sensing (QS) system in S. aureus through regulating the auto inducer 2 (AI-2) activity, which has been reported to be negative for biofilm formation in S. aureus. These above properties could be applied in developing surfactin as a potential pre-coating agent on material surfaces to prevent S. aureus biofilm formation.

Preparation of chitosan/curdlan/carboxymethyl cellulose blended film and its characterization.

In this study, to improve the thermal and mechanical properties of chitosan films, a chitosan/curdlan/carboxymethyl cellulose (CS/CD/CMC) ternary blended film was prepared and characterized. To prepare a uniform CS/CD/CMC ternary blended film, an effective method of blending CD with other materials was established as the following conditions: the ternary solution temperature was maintained at 60 °C, and the pH was controlled in the range from 12 to 4. Compared to the pure chitosan, the CS/CD/CMC blended films exhibited better mechanical properties, permeability, and thermal stability. In addition, visible light properties of the ternary blending film were improved. Scanning electron microscope and Fourier transform-infrared spectroscopy analyses indicated good compatibility among the CS, CD and CMC, which led to a corresponding improvement in the properties owing to interactions among the three components in the blending process. So, an effective method of blending CD with CS and CMC was established, and the blending film has good thermal and mechanical properties.

Consensus design for improved thermostability of lipoxygenase from Anabaena sp. PCC 7120.

BACKGROUND: Lipoxygenase (LOX) from Anabaena sp. PCC 7120 (Ana-rLOX) offers important applications in the food industry, especially for improving aroma and dough rheological properties. However, industrial applications of LOXs have been limited by their poor thermostability. Herein, we report a bioinformatics-based consensus concept approach for the engineering of thermostable Ana-rLOX. RESULTS: A series of mutations (N130D, G260A, S437T, N130D/G260Q, N130D/S437Y) showed higher thermostability and activity than the wild-type enzyme. Thus, N130D/G260Q exhibited a 6.6-fold increase in half-life and 2.45 °C increase in unfolding temperature; N130D/S437Y showed a 10 °C increase in optimal temperature. The secondary structure did not change much that contributed to improved thermostability were investigated in detail using circular dichroism. Homology modeling suggested that enhanced thermostability and specific activity may result from favorable hydrophobic interactions. CONCLUSIONS: A series of mutations were achieved, showing higher thermostability and activity than the wild-type enzyme by semi-rational mutagenesis with limited structure information. Our findings provide important new insights into molecular modifications aimed at improving Ana-rLOX thermostability and activity.

Module and individual domain deletions of NRPS to produce plipastatin derivatives in Bacillus subtilis.

BACKGROUND: Plipastatin, an antifungal lipopeptide, is synthesized by a non-ribosomal peptide synthetase (NRPS) in Bacillus subtilis. However, little information is available on the combinatorial biosynthesis strategies applied in plipastatin biosynthetic pathway. In this study, we applied module or individual domain deletion strategies to engineer the plipastatin biosynthetic pathway, and investigated the effect of deletions on the plipastatin assembly line, as well as revealed the synthetic patterns of novel lipopeptides. RESULTS: Module deletion inactivated the entire enzyme complex, whereas individual domain (A/T domain) deletion within module 7 truncated the assembly line, resulting in truncated linear hexapeptides (C16~17ß-OHFA-Glu-Orn-Tyr-Thr-Glu-Ala/Val). Interestingly, within the module 6 catalytic unit, the effect of thiolation domain deletion differed from that of adenylation deletion. Absence of the T6-domain resulted in a nonproductive strain, whereas deletion of the A6-domain resulted in multiple assembly lines via module-skipping mechanism, generating three novel types of plipastatin derivatives, pentapeptides (C16~17ß-OHFA-Glu-Orn-Tyr-Thr-Glu), hexapeptides (C16~17ß-OHFA-Glu-Orn-Tyr-Thr-Glu-Ile), and octapeptides (C16~17ß-OHFA-Glu-Orn-Tyr-Thr-Glu-Gln-Tyr-Ile). CONCLUSIONS: Notably, a unique module-skipping process occurred following deletion of the A6-domain, which has not been previously reported for engineered NRPS systems. This finding provides new insight into the lipopeptides engineering. It is of significant importance for combinatorial approaches and should be taken into consideration in engineering non-ribosomal peptide biosynthetic pathways for generating novel lipopeptides.

The antibacterial activity of LI-F type peptide against methicillin-resistant Staphylococcus aureus (MRSA) in vitro and inhibition of infections in murine scalded epidermis.

LI-F type peptides are a family of cyclic lipodepsipeptide antibiotics isolated from Paenibacillus polymyxa and display potent activities against positive bacteria including methicillin-resistant S. aureus (MRSA). In this study, we investigated the mechanism of action of LI-F type peptide AMP-jsa9 against a MRSA (S. aureus CICC10790), which is resistant to ciprofloxacin, gentamicin, kanamycin, chloramphenicol, methicillin, and tetracycline. It was found that AMP-jsa9 mainly targets the cell membrane of MRSA and is able to inhibit biofilm formation through killing planktonic bacteria cells. Moreover, AMP-jsa9 can bind to DNA in vitro, which represents another pathway for the action on MRSA. Furthermore, in vivo treatment of scalded mice with AMP-jsa9 resulted in inhibiting MRSA infections and healing of the scalded wound. In addition, it was demonstrated that AMP-jsa9 can effectively inhibit MRSA infections in scalded murine epidermis and that inflammatory cytokines including IL-8, IL-6, tumor necrosis factor alpha (TNF-α), and monocyte chemotactic factor-1 (MCP-1) were reduced; moreover, both protein and gene expression levels of vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (e-NOS) were enhanced, which promote neovascularization and proliferation of new granulation tissue.

Expression, purification, and characterization of a novel acidic Lipoxygenase from Myxococcus xanthus.

The gene encoding a novel acidic lipoxygenase from Myxococcus xanthus DK1622 (accession: WP_011551853.1) was cloned into vector pET-28a and expressed in Escherichia coli BL21(DE3). The recombinant enzyme (rMxLOX), with a molecular weight of approximately 80 kDa, was purified to homogeneity using one-step nickel-affinity chromatography and showed an activity of 5.6 × 104 U/mg. The optimum pH and temperature for rMxLOX activity were found to be 3.0 and 30 °C, respectively. Purified rMxLOX exhibited activity towards linoleic acid and arachidonic acid as substrates, with linoleic acid being the better substrate (Km and kcat values of 0.048 mM and 13.3/s, respectively). The synthetic dye aniline blue was decolorized 69.7 ± 3.5%, following incubation with rMxLOX for 35 min. These results reveal the potential for the use of rMxLOX in the pulp, textile, and wastewater treatment industries.

Identification of novel surfactin derivatives from NRPS modification of Bacillus subtilis and its antifungal activity against Fusarium moniliforme.

BACKGROUND: Bacillus subtilis strain PB2-L1 produces the lipopeptide surfactin, a highly potent biosurfactant synthesized by a large multimodular nonribosomal peptide synthetase (NRPS). In the present study, the modules SrfA-A-Leu, SrfA-B-Asp, and SrfA-B-Leu from surfactin NRPS in B. subtilis BP2-L1 were successfully knocked-out using a temperature-sensitive plasmid, pKS2-mediated-based, homologous, recombination method. RESULTS: Three novel surfactin products were produced, individually lacking amino acid Leu-3, Asp-5, or Leu-6. These surfactins were detected, isolated, and characterized by HPLC and LC-FTICR-MS/MS. In comparison with native surfactin, [∆Leu(3)]surfactin and [∆Leu(6)]surfactin showed evidence of reduced toxicity, while [∆Asp(5)]surfactin showed stronger inhibition than native surfactin against B. pumilus and Micrococcus luteus. These results showed that the minimum inhibitory concentration of [∆Leu(6)]surfactin for Fusarium moniliforme was 50 µg/mL, such that [∆Leu(6)]surfactin could lead to mycelium projection, cell damage, and leakage of nucleic acids and protein. These factors all contributed to stimulating apoptosis in F. moniliforme. CONCLUSIONS: The present results revealed that [∆Leu(6)]surfactin showed a significant antifungal activity against F. moniliforme and might successfully be employed to control fungal food contamination and improve food safety.

Expression of a novel bacteriocin-the plantaricin Pln1-in Escherichia coli and its functional analysis.

A potential bacteriocin gene was isolated from 18575 ORFs by bioinformatics methods. It was named pln1, and cloned into pET32a. Then, it was expressed as a thioredoxin-Pln1 fusion protein in Escherichia coli BL21 (DE3). The fusion protein was purified by Ni-NTA, and thioredoxin was removed by enterokinase. Finally, Pln1 was purified using a cation affinity column. The yields of fused and cleaved Pln1 peptides were 100-110 mg/l and 9-11 mg/l, respectively. Pln1 was stable in an acidic environment and at temperatures below 60 °C, but was easily degraded under alkaline conditions and by protease treatment. The cleaved and purified Pln1 showed strong antimicrobial activity against gram-positive bacteria such as Micrococcus luteus CMCC 63202, Staphylococcus epidermidis, Lactococcus lactis NZ3900, Lactobacillus paracasei CICC 20241, and Listeria innocua CICC 10417. In particular, Pln1 had a better activity against methicillin-resistant S. epidermidis (MRSE) than nisin, thereby offering an attractive approach to counter bacterial antibiotic resistance.

Effects of recombinant lipoxygenase on the rheological properties of dough and the quality of noodles.

BACKGROUND: The effects of purified recombinant lipoxygenase (ana-rLOX) on the rheological properties of dough and the quality of noodles made from wheat flour with low protein content (Yanmai 15) were studied. RESULTS: The addition of ana-rLOX increased dough stability time, decreased the degree of softening within 12 min, enhanced the resistance to extension, and increased the extensibility with 135 min of resting time. The mechanical spectra of the dough showed an increase in both storage modulus (G') and loss modulus (G″) with increasing ana-rLOX levels. The L(*) values of the noodle sheets increased by 2.34 compared with the control after storing for 1 h at room temperature. The textural parameters of noodles improved after ana-rLOX addition, including hardness, gumminess, chewiness and springiness. The wheat flour treated with the ana-rLOX had a higher cooking yield and lower cooking loss for the resulting noodles. The scanning electron microscopy results revealed that gluten was formed in the noodle samples that were treated with ana-rLOX. CONCLUSION: In this study, ana-rLOX was applied to noodles during the noodle-making process, and both dough rheological characteristics and noodle quality were improved. © 2015 Society of Chemical Industry.

Engineering of a thermostable ß-1,3-1,4-glucanase from Bacillus altitudinis YC-9 to improve its catalytic efficiency.

BACKGROUND: Error-prone polymerase chain reaction (PCR) is frequently used in directed evolution of enzymes to modify their quality. In this study, error-prone PCR was used to improve the catalytic efficiency of ß-1,3-1,4-glucanase from Bacillus altitudinis YC-9. RESULTS: By screening, the mutant Glu-3060 with higher activity was selected among 5000 transformants. After induction with isopropyl ß-D-1-thiogalactopyranoside (IPTG), the activity of the mutant Glu-3060 reached 474.6 U mL(-1), resulting in a 48.6% increment of the parent enzyme activity. Research on the characterization of the mutated enzyme showed the optimal pH of the mutated enzyme to be 5.0, which is lower than the parent enzyme, but thermal stability was almost the same between them. Sequence analysis of the mutated enzyme revealed that three amino acids were changed compared with the parent enzyme, including K142N, Q203L and N214D. CONCLUSION: The three-dimensional structure predicted by SWISS-MODEL of the mutated enzyme Glu-3060 showed that the substitution of three amino acids had an effect on the catalytic activity, stability and optimal pH of the enzyme, through changing the charge properties or electron density, forming secondary keys, the acidity of the amino acids and the side chain group. The sum effects of all the factors were increased activity of the mutated enzyme and decreased optimal pH, while the same thermostability was maintained, thereby increasing the suitability of the enzyme for industrial use.