Enhancement of the yield of poly (ethylene terephthalate) hydrolase production using cell membrane protection strategy.

Biodegradation, particularly via enzymatic degradation, has emerged as an efficient and eco-friendly solution for Poly (ethylene terephthalate) (PET) pollution. The production of PET hydrolases plays a role in the large-scale enzymatic degradation. However, an effective variant, 4Mz, derived from Thermobifida fusca cutinase (Tfu_0883), was previously associated with a significant reduction in yield when compared to the wild-type enzyme.

High-efficient preparation of β-nicotinamide mononucleotides by crude enzymes cascade catalytic reaction.

β-nicotinamide mononucleotide (β-NMN) is a key precursor of nicotinamide adenine dinucleotide, and becomes attractive in the nutrition and health care fields, but its enzymatic synthesis is expensive. In this study, a six-enzyme cascade catalytic system was constructed to produce β-NMN. Using D-ribose and nicotinamide as substrates, the β-NMN yield reached 97.

Genome-Wide CRISPRi Screening of Key Genes for Recombinant Protein Expression in Bacillus Subtilis.

Bacillus subtilis is an industrially important microorganism that is often used as a microbial cell factory for the production of recombinant proteins due to its food safety, rapid growth, and powerful secretory capacity. However, the lack of data on functional genes related to recombinant protein production has hindered the further development of B. subtilis cell factories.

Enzymatic treatment to improve permeability and quality of cherry tomatoes for production of dried products.

Background: Cherry tomatoes are nutritious and favored by consumers. Processing them into dried cherry tomatoes can prolong their storage life and improve their flavor. The pretreatment of tomato pericarp is crucial for the subsequent processing.

High-Level Production of Lacto--neotetraose in by Stepwise Optimization of the Biosynthetic Pathway.

Lacto--neotetraose (LNnT), an abundant human milk oligosaccharide (HMO), has been approved as a novel functional additive for infant formulas. Therefore, LNnT biosynthesis has attracted extensive attention. Here, a high LNnT-producing, low lacto--triose II (LNT II)-residue strain was constructed.

High-Yield Synthesis of 2'-Fucosyllactose from Glycerol and Glucose in Engineered .

2'-Fucosyllactose (2'-FL) is vital for the growth and development of newborns. In this study, we developed a synthesis pathway for 2'-FL in BL21 (DE3). Then, we optimized the solubility of α-1,2-fucosyltransferase, thereby enhancing the production yield of 2'-FL.

Conformational Switch of the 250s Loop Enables the Efficient Transglycosylation in GH Family 77.

Amylomaltases (AMs) play important roles in glycogen and maltose metabolism. However, the molecular mechanism is elusive. Here, we investigated the conformational dynamics of the 250s loop and catalytic mechanism of AM using path-metadynamics and QM/MM MD simulations.

Differentiated digestion resistance and physicochemical properties of linear and α-1,2/α-1,3 branched isomaltodextrins prepared by 4,6-α-glucanotransferase and branching sucrases.

Isomaltodextrins (IMDs) are starch-based dietary fibers (DF) prepared enzymatically, which show great potential as a functional food ingredient. In this study, a series of novel IMDs with diverse structures were generated by 4,6-α-glucanotransferase GtfBΔN from Limosilactobacillus fermentum NCC 3057, combined with two α-1,2 and α-1,3 branching sucrases. Results indicated that α-1,2 and α-1,3 branching significantly improved the DF contents of α-1,6 linear products up to 60.

Structural characterization of slow digestion dextrin synthesized by a combination of α-glucosidase and cyclodextrin glucosyltransferase and its prebiotic potential on the gut microbiota in vitro.

Starch-based dietary fibers are at the forefront of functional ingredient research. In this study, a novel water-soluble slow digestion dextrin (SDD) was synthesized by synergy of α-glucosidase and cyclodextrin glucosyltransferase and characterized. Results showed that SDD exhibited high solubility, low viscosity, and resistance to digestive enzymes, and also showed an increased dietary fiber content of 45.

Mechanistic Insights into How the Protonation State of D234 Dictates the Reactivity in β--Acetylhexosaminidase.

β--Acetylhexosaminidases (HEXs) play important roles in human diseases and the biosynthesis of human milk oligosaccharides. Despite extensive research, the catalytic mechanism of these enzymes remains largely unexplored. In this study, we employed quantum mechanics/molecular mechanics metadynamics to investigate the molecular mechanism of HEX (HEX), which has shed light on the transition state structures and conformational pathways of this enzyme.

Enhancing 2-O-α-D-glucopyranosyl-L-ascorbic acid synthesis by weakening the acceptor specificity of CGTase toward glucose and maltose.

2-O-α-D-glucopyranosyl-L-ascorbic acid (AA-2G) is a stable derivative of L-ascorbic acid (L-AA), which has been widely used in food and cosmetics industries. Sugar molecules, such as glucose and maltose produced by cyclodextrin glycosyltransferase (CGTase) during AA-2G synthesis may compete with L-AA as the acceptors, resulting in low AA-2G yield. Multiple sequence alignment combined with structural simulation analysis indicated that residues at positions 191 and 255 of CGTase may be responsible for the difference in substrate specificity.

Multiple approaches of loop region modification for thermostability improvement of 4,6-α-glucanotransferase from Limosilactobacillus fermentum NCC 3057.

4,6-α-glucanotransferase (4,6-α-GT), as a member of the glycoside hydrolase 70 (GH70) family, converts starch/maltooligosaccharides into α,1-6 bond-containing α-glucan and possesses potential applications in food, medical and related industries but does not satisfy the high-temperature requirement due to its poor thermostability. In this study, a 4,6-α-GT (ΔGtfB) from Limosilactobacillus fermentum NCC 3057 was used as a model enzyme to improve its thermostability. The loops of ΔGtfB as the target region were optimized using directed evolution, sequence alignment, and computer-aided design.

Producing 2-O-α-D-glucopyranosyl-L-ascorbic acid by modified cyclodextrin glucosyltransferase and isoamylase.

In this study, site saturation mutagenesis was performed on the - 3 (R44, D86, S90, and D192) and - 6 subsite (Y163, G175, G176, and N189) of Bacillus stearothermophilus NO2 cyclodextrin glucosyltransferase to enhance its specificity for the donor substrate maltodextrin for 2-O-α-D-glucopyranosyl-L-ascorbic acid (AA-2G) preparation. The AA-2G yields produced by the mutants S90D, G176H, and S90D/G176H were 181, 171, and 185 g/L, respectively. Our previous study found that the mutant K228R/M230L also increased the AA-2G yield.

Oxidative degradation of pre-oxidated polystyrene plastics by dye decolorizing peroxidases from Thermomonospora curvata and Nostocaceae.

Biodegradation of PS has attracted lots of public attentions due to its environmental friendliness. However, no specific PS degrading enzyme has been identified yet. Dye decolorizing peroxidases (DyPs) are heme-containing peroxidases named for the ability to degrade a variety of organic dyes.

A Single Hydrogen Bond Controls the Selectivity of Transglycosylation vs Hydrolysis in Family 13 Glycoside Hydrolases.

Converting glycoside hydrolases (GHs) from hydrolytic to synthetic enzymes via transglycosylation is a long-standing goal for the biosynthesis of complex carbohydrates. However, the molecular determinants for the selectivity of transglycosylation (T) vs hydrolysis (H) are still not fully unraveled. Herein, we show experimentally that mutation of one active site residue can switch the enzyme activity between hydrolysis and transglycosylation in two highly homologous GHs.

Trehalose promotes high-level heterologous expression of 4,6-α-glucanotransferase GtfR2 in Escherichia coli and mechanistic analysis.

4,6-α-Glucanotransferases (4,6-α-GTs) hold great potential for applications in the food and medical industries because of their efficient transglycosylation ability. However, it is relatively difficult to achieve high soluble expression because of their high molecular weight and multidomain nature. In this study, 4,6-α-GT of Burkholderia sp.

Directed Mutation of Two Key Amino Acid Residues Alters the Product Structure of the New 4,6-α-Glucanotransferase from .

4,6-α-Glucanotransferases (4,6-α-GTs) convert amylose V into two types of differently structured products: a linear product connected by continuous α,1 → 6 bonds, such as isomalto/malto-polysaccharide (IMMP), and a highly branched product connected by alternating α,1 → 4 and α,1 → 6 bonds, such as reuteran-like polysaccharide (RLP). The synthesis process of 4,6-α-GT products is unclear, and exploring this process is significant for producing dietary fibers with potential applications. This study identified and expressed sp.

Enhancing Extracellular Pullulanase Production in Bacillus subtilis Through dltB Disruption and Signal Peptide Optimization.

Bacillus subtilis has many attributes that make it a popular host for recombinant protein production. Although its protein production ability has been enhanced through protease gene disruption, residual proteases like quality control HtrA and HtrB can limit protein yield by degrading inadequately folded proteins present during overexpression. In this study, two strategies were employed to increase production of industrial enzyme pullulanase: enhancing extracellular pullulanase folding and optimizing its signal peptide.

Enhanced extracellular α-amylase production in Brevibacillus choshinensis by optimizing extracellular degradation and folding environment.

A strategy for optimizing the extracellular degradation and folding environment of Brevibacillus choshinensis has been used to enhance the extracellular production of recombinant α-amylase. First, a gene (bcp) encoding an extracellular protease and another encoding an extracellular chaperone (prsC) were identified in the genome of B. choshinensis HPD31-SP3.

Microbial starch debranching enzymes: Developments and applications.

Starch debranching enzymes (SDBEs) hydrolyze the α-1,6 glycosidic bonds in polysaccharides such as starch, amylopectin, pullulan and glycogen. SDBEs are also important enzymes for the preparation of sugar syrup, resistant starch and cyclodextrin. As the synergistic catalysis of SDBEs and other starch-acting hydrolases can effectively improve the raw material utilization and production efficiency during starch processing steps such as saccharification and modification, they have attracted substantial research interest in the past decades.

Effect of Leu on Disproportionation and Hydrolysis Activity in Bacillus stearothermophilus NO2 Cyclodextrin Glucosyltransferase.

The disproportionation activity of cyclodextrin glucosyltransferase (CGTase; EC 2.4.1.

Enhanced Production of Soluble α-Amylase in through Chaperone Co-Expression, Heat Treatment and Fermentation Optimization.

α-amylase can hydrolyze α-1,4 linkages in starch and related carbohydrates under hyperthermophilic condition (~ 100°C), showing great potential in a wide range of industrial applications, while its relatively low productivity from heterologous hosts has limited the industrial applications. , a gram-positive bacterium, has been widely used in industrial production for its non-pathogenic and powerful secretory characteristics. This study was conducted to increase production of α-amylase in through three strategies.