Enhanced catalytic performance and pH stability of Streptomyces Laccase Y230R and its degradation of malachite green.

The escalating threat of malachite green (MG) pollution poses significant risks to ecosystems. Saturation mutation targeting Tyr230 of small laccase (SLAC) from Streptomyces coelicolor yielded Y230R, exhibiting a remarkable 104 % increase in specific activity. Notably, this mutation achieved dual enhancements in both activity and pH stability. Molecular dynamics simulation revealed higher structural stability of Y230R compared to wild-type (WT) across varying pH levels. The increased count of hydrogen bonds in Y230R compared to WT may be contribute to its stability. Y230R demonstrated superior catalytic efficiency (67.0 %) in MG decolorization, maintaining over 90 % activity after 30 min incubation in MG solution (500 mg/L), highlighting enhanced tolerance compared to WT. Molecular docking analysis attributed the differential catalytic effects on MG and ABTS to structural disparities and hydrogen bonding. Y230R stands as a promising composite mutant for future laccase engineering and industrial applications.

Impacts of transglutaminase on the processing and digestion characteristics of glutinous rice flour: Insight of the interactions between enzymic crossing-linked protein and starch.

Glutinous rice is extensively consumed due to its nutritious content and wonderful flavor. However, glutinous rice flour has a high glycemic index, and the storage deterioration of sweet dumplingsissevere. Transglutaminase (TG) was used to cross-link glutinous rice protein and improve the characteristics of glutinous rice products. The findings demonstrated that TG significantly catalysed protein cross-linking to form a dense protein network, reduced the viscosity of glutinous rice paste and improved the thermal stability. The protein network may physically block the access of starch granules to digestive enzymes to lower the digestion rate of starch, and attenuate the damage of ice crystal molecules to the starch structure to improve the freezing stability of starch gels. The cracking rate and water loss of sweet dumplings prepared using glutinous rice flour with TG treated for 60 min reduced significantly. In conclusion, this study broadened the application of TG in starch products.

Engineering Site 228 of Streptomyces coelicolor Laccase for Optimizing Catalytic Activity.

Malachite green (MG) poses a formidable threat to ecosystems and human health. Laccase emerges as a promising candidate for MG degradation, prompting an investigation into the catalytic activity modulation of a small laccase (SLAC) from Streptomyces coelicolor, with a focus on amino acid position 228. Through saturation mutagenesis, five mutants with a 50% increase in the specific activity were generated. Characterization revealed notable properties, Km of E228F was 8.8% of the wild type (WT), and E288T exhibited a 133% kcat compared to WT. Structural analyses indicated improved hydrophobicity and electrostatic potential on the mutants' surfaces, with the stable E228F-ABTS complex exhibiting reduced flexibility, possibly contributing to the observed decrease in turnover rate. Mutants demonstrated enhanced MG decolorization, particularly E228G. Site 228 acts as a crucial functional control switch, suggesting its potential role in SLAC engineering. This study provides insights into laccase modulation and offers promising avenues for enzymatic bioremediation applications.

Enhancing the Physicochemical Attributes of Dough and Noodles through the Incorporation of Bacillus vallismortis Laccase.

This investigation examined how the Bacillus vallismortis laccase (rBVL-MRL522) influenced the physicochemical characteristics, structural attributes, and functional capabilities of both dough and noodles. Incorporating rBVL-MRL522 (1 U/g) did not lead to a substantial change in the water absorption of wheat flour. However, the introduction of rBVL-MRL522 caused a significant elongation in the formation time of wheat flour dough, extending it by 88.9%, and also resulted in a 50% increase in the stabilization duration of wheat flour dough. Furthermore, adding rBVL-MRL522 led to a proportional rise in both the elastic and viscous moduli (G'' of the dough, signifying that r-BVL (rBVL-MRL522) has a beneficial effect on the gluten strength of the dough. Integrating rBVL-MRL522 promoted the consolidation of the gluten-based cross-linked structure within the dough, decreasing the size of starch particles and, more evenly, the dispersion of these starch particles. In the noodle processing, adding rBVL-MRL522 at a rate of 1 U/g raised the L* value of the noodles by 2.34 units compared to the noodles prepared without the inclusion of rBVL-MRL522. Using a greater amount of rBVL-MRL522 (2 U/g) substantially increased the hardness of the noodles by 51.31%. Additionally, rBVL-MRL522 showed a noteworthy enhancement in the elasticity, cohesiveness, and chewiness of the noodles. In conclusion, rBVL-MRL522 promoted the cross-linking gluten, leading to a more extensive and condensed three-dimensional network structure in raw and cooked noodles. As a result, this study offers valuable insights into the environmentally friendly processing of dough and associated products.

Study of alkali-soluble curdlan/bacterial cellulose/cinnamon essential oil blend films with enhanced mechanical properties.

In response to the growing demand for biodegraded film with high mechanical properties for complex preservation application scenarios, we developed a curdlan (CD) blended films with exceptional mechanical strength through an alkali dissolution method. Notably, the alkali-soluble CD film exhibited five-fold increase in tensile strength (TS) when compared to its water-soluble counterpart. Furthermore, the inclusion of 2 % bacterial cellulose (BC) resulted in a significant 41.1 % augmentation of the film's TS. Thermal stability improvements were observed through differential scanning calorimetry (DSC) analysis and thermogravimetric analysis (TGA). X-ray diffraction (XRD) and Fourier-transform infrared spectroscopy (FTIR) results provided insights into alterations in film crystallinity and intermolecular interactions. Specifically, the incorporation of 10 % CEO led to an additional improvement in TS. Our experimental investigations involving the packaging of chilled fresh meat with these blended films unveiled their capacity to effectively inhibit microorganism growth, maintain meat color stability, delay protein decomposition and fat oxidation, and extend the storage time up to 9 days. Our study offers a promising solution for food packaging, emphasizing the development of a high-strength degradable CD/BC/CEO blended film, which holds potential for extending the shelf life of food products.

Enhancement of Antioxidant Property of N-Carboxymethyl Chitosan and Its Application in Strawberry Preservation.

Bio-enzymatic grafting phenolic acid to chitosan derivative is an efficient and environmentally friendly molecular synthesis technology. In the present study, N-carboxymethyl chitosan (CMCS) was grafted with gallic acid (GA) using recombinant bacterial laccase from Streptomyces coelicolor as a catalyst. GA and CMCS were successfully grafted as determined by measuring amino acid content, Fourier transform infrared (FTIR) spectroscopy and ultraviolet-visible (UV-Vis) spectroscopy. Then, the effect of GA-g-CMCS coating on the freshness of strawberries at 20 ± 2 °C was explored. The physiological and biochemical quality indicators of strawberries during storage were monitored. The 1.5% GA-g-CMCS coating helped to protect the antioxidant properties and nutrients of strawberries and extend the shelf life. Specifically, it reduced the weight loss of strawberries during preservation (originally 12.7%) to 8.4%, maintained titratable acidity content (TA) residuals above 60% and reduced decay rate from 36.7% to 8.9%. As a bioactive compound, GA-g-CMCS has the potential to become an emerging food packing method. These results provide a theoretical basis and reference method for the subsequent synthesis and application of CMCS derivatives.

Degradation of Aflatoxin B1 by recombinant laccase extracellular produced from Escherichia coli.

Bioenzymatic degradation of aflatoxin B1 (AFB1) is a safe, efficient and environmentally friendly detoxification technology. In this work, AFB1 was successfully degraded by recombinant laccase (fmb-rL103) in the absence of a mediator. The laccase gene was cloned from Bacillus vallismortis fmb-103, and was expressed in heterologous host Escherichia coli after codon optimization. The extracellular production of fmb-rL103 could be induced by adding methanol (6 %, v/v), and the maximum yield was 1545.6 U/L. In the 10 L bioreactor, the extracellular yield increased to 50,950.6 U/L after 20 h of induction, accounting for three quarters of the total yield. The mechanism of methanol-induced extracellular secretion was further studied by measuring acetate content, lac103 gene expression and cell membrane permeability. Furthermore, we explored the biochemical properties of fmb-rL103 and its degradation conditions on AFB1. The degradation efficiency increased constantly with increase in incubation pH and temperature, and exceeded 60 % at pH 7.0 and 37 °C. This work provides new insight into developing the large-scale production of laccase and its application to degrade AFB1.

Preparation of a novel curdlan/bacterial cellulose/cinnamon essential oil blending film for food packaging application.

With the increasing attention to food preservation and environmental safety, there is great pressing demand to explore novel edible and environment-friendly food packaging films. In the present study, a new kind natural curdlan (CD) film was developed with the addition of bacterial cellulose (BC) and cinnamon essential oil (CEO) at 2% and 10% (w/w) amounts, with regard to improve mechanical properties and investigate potential food applications. Our results showed that the tensile strength, the crystallinity and the thermal stability of the CD/BC blending film were improved, while the water vapor permeability, moisture content and the lightness were reduced. Moreover, the CEO addition to the CD/BC film further increased the barrier properties and also mechanical properties. The results of FTIR and XRD were applied for analyzing the potential interactions of the film matrix. Finally, addition of CEO endowed the blending films with good antibacterial activity and antioxidant capacity, which could effectively inhibit the bacterial growth and the lipid oxidation of chilled chicken during the preservative period. Thus, this work demonstrates that the novel CD/BC/CEO blending film with improved mechanical and barrier properties can be of great potential for developing food packaging material for promising applications.

Preparation and characterization of curdlan/nanocellulose blended film and its application to chilled meat preservation.

In this study, to improve the mechanical and thermal properties of curdlan film, a curdlan/nanocellulose (NC) blended film was prepared and characterized for the first time. NC was successfully prepared from microcrystalline cellulose (MCC) with NaOH/urea treatment. The particle size of NC was observed to be 70-140 nm by cryo-electron microscope (cryo-EM). The blended film was prepared by adding the NC to curdlan solution. The tensile strength (TS) of the blended film reached the maximum value of 38.6 MPa, and the elongation at break (EB) was 40%. The DSC curve showed that the heat absorption peak of the film was 240 °C, indicating that the blended film has good temperature stability. Additionally, some other film properties were also improved, including gas barrier properties and transparency. Obvious morphological and molecular differences between the blended film and the pure curdlan film were discovered by SEM and FTIR analysis. Finally, the blended film was used for the preservation of chilled meat and extended the storage time of meat to 12 days. These results provided a theoretical basis for future application and development of biodegradable film.