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D-tagatose, a low-calorie rare sugar, has significant potential in food, medicine, cosmetics, and other industries owing to its high application value and market potential. In this study, Escherichia coli BL21 was used as the starting strain to express the β-galactosidase (β-Gal) gene—BgaB—derived from Bacillus stearothermophilus and the L-arabinose isomerase (L-AI) gene—araA—derived from Thermus sp., yielding the genetically engineered strains E. coli BL21-pET28a-BgaB and E. coli BL21-pET28a-araA. These strains synthesized D-tagatose using β-Gal and L-AI with a conversion rate of 23.73%. Based on this, we constructed a multienzyme cascade pathway comprising β-Gal, L-AI, glucose isomerase (GI), fructose kinase (FK), D-tagatose-bisphosphate aldolase (GatZ), polyphosphate kinase (PPK), and phosphatase (PGP), further enhancing D-tagatose biosynthesis. This multienzyme approach improved the conversion of the intermediate product D-glucose to D-tagatose by 3.84% compared with the dual-enzyme system. Thus, our study provides a theoretical basis and technical support for the industrial production of D-tagatose. [ABSTRACT FROM AUTHOR]

D-tagatose is a rare, naturally occurring low-calorie hexose, with a sweetness of 92% sucrose but only 1/3 of the calories. It has beneficial functions in lowering blood sugar, controlling obesity, preventing dental caries, and improving intestinal flora. In recent years, biotechnological routes to D-tagatose production from renewable raw materials have been regarded as very promising approaches. In this review, we provide an overview of the properties and applications of D-tagatose, with a focus on the current developments in the production of D-tagatose using enzymatic transformation and whole-cell catalytic synthesis. The biosynthetic pathways and the types and characteristics of the catalytic enzymes involved have been summarized, providing a reference for the design of D-tagatose synthesis pathways. We also expect that rapid developments in the fields of systems biology and synthetic biology will accelerate protein and metabolic engineering for microbial D-tagatose production in the future. [ABSTRACT FROM AUTHOR]

D-tagatose is a valuable rare sugar with potential health benefits such as antiobesity, low-calorie, prebiotic, and anticancer. However, its production is mainly depending on chemical or enzymatic catalysis. Herein, a cobalt-based metal–organic framework (MOF) was developed at room temperature in an aqueous system using a self-assembly method. The L-arabinose isomerase (L-AI) was immobilized into this unique MOF by an in situ encapsulation process. The morphology and structural aspects of the MOF preparations were characterized by different analytical techniques such as scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX), confocal laser scanning microscopy (CLSM), Fourier transform infrared spectroscopy (FT-IR), and X-Ray diffraction (XRD). Moreover, thermogravimetric analysis (TGA) suggested the high thermal stability of the L-AI@MOF. Significantly, the immobilized catalyst exhibited enhanced catalytic efficiency (kcat/Km) of 3.22 mM−1 s−1 and improved turnover number (kcat) of 57.32 s−1. The L-AI@MOF efficiently catalyzes the synthesis of D-tagatose from D-galactose up to the equilibrium level (~ 50%) of isomerization in heterogeneous catalysis. Interestingly, L-AI@MOF was found stable and reusable for more than five cycles without the requirement of additional metal ions during catalysis. Thus, L-AI stabilized in the MOF system demonstrated a higher catalytic activity and potential guidance for the sustainable synthesis of rare sugar D-tagatose. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Food Science & Biotechnology is the property of Journal of Food Science & Biotechnology Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Background: stevia and D-tagatose have shown a reduction in total calorie and carbohydrate intake as a substitute for sucrose, demonstrating a stabilizing effect on pH and bacterial proliferation. Objective: to evaluate the effect of D-tagatose, stevia and sucrose on salivary pH and bacterial activity in odontology students. Methodology: a controlled study of parallel and randomized groups with a single blind, whose sample considered three groups subjected to a mouthwash of D-tagatose (n = 10), stevia (n = 10) and sucrose (n = 10). These solutions were administered over 1 minute in a single 6.4 % concentrated dose. Data collection and analysis considered the recording of salivary pH 5 min before exposure to the sweetener, immediately after expulsion of the mouthwash and 15 min later, 30 min, 45 min and 48 hours. The counting of the final number of colony-forming units per mL (CFU/mL) was counted using the salivary samples obtained immediately after exposure of the sweetener together with the sample obtained 30 minutes later, with the cultures performed on agar plates. Results: D-tagatose, stevia and sucrose presented significant differences in total CFU/mL at 30 minutes (p < 0.001), while salivary pH showed significant differences at 48 hours after administration (p < 0.001). Conclusion: D-tagatose, stevia and sucrose present significant differences in total CFU/mL and salivary pH, these findings being a possible indication of a partial inhibitory effect on bacterial metabolism. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Dalian Polytechnic University is the property of Journal of Dalian Polytechnic University Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

D-tagatose is a rare, naturally occurring low-calorie hexose, with a sweetness of 92% sucrose but only 1/3 of the calories. It has beneficial functions in lowering blood sugar, controlling obesity, preventing dental caries, and improving intestinal flora. In recent years, biotechnological routes to D-tagatose production from renewable raw materials have been regarded as very promising approaches. In this review, we provide an overview of the properties and applications of D-tagatose, with a focus on the current developments in the production of D-tagatose using enzymatic transformation and whole-cell catalytic synthesis. The biosynthetic pathways and the types and characteristics of the catalytic enzymes involved have been summarized, providing a reference for the design of D-tagatose synthesis pathways. We also expect that rapid developments in the fields of systems biology and synthetic biology will accelerate protein and metabolic engineering for microbial D-tagatose production in the future.

L-Arabinose isomerase (L-AI) has been commonly used as an efficient biocatalyst to produce D-tagatose via the isomerization of D-galactose. However, it remains a significant challenge to efficiently synthesize D-tagatose using the native (wild type) L-AI at an industrial scale. Hence, it is extremely urgent to redesign L-AI to improve its catalytic efficiency towards D-galactose, and herein a structure-based molecular modification of Lactobacillus plantarum CY6 L-AI (LpAI) was performed. Among the engineered LpAI, both F118M and F279I mutants showed an increased D-galactose isomerization activity. Particularly, the specific activity of double mutant F118M/F279I towards D-galactose was increased by 210.1% compared to that of the wild type LpAI (WT). Besides the catalytic activity, the substrate preference of F118M/F279I was also largely changed from L-arabinose to D-galactose. In the enzymatic production of D-tagatose, the yield and conversion ratio of F118M/F279I were increased by 81.2% and 79.6%, respectively, compared to that of WT. Furthermore, the D-tagatose production of whole cells expressing F118M/F279I displayed about 2-fold higher than that of WT cell. These results revealed that the designed site-directed mutagenesis is useful for improving the catalytic efficiency of LpAI towards D-galactose. [ABSTRACT FROM AUTHOR]

D-allulose, D-sorbose and D-tagatose are D-fructose isomers that are called rare sugars. These rare sugars have been studied intensively in terms of biological production and food application as well as physiological effects. There are limited papers with regard to the transporters mediating the intestinal absorption of these rare sugars. We examined whether these rare sugars are absorbed via sodium-dependent glucose cotransporter 1 (SGLT1) as well as via GLUT type 5 (GLUT5) using rats. High-fructose diet fed rats, which express more intestinal GLUT5, exhibited significantly higher peripheral concentrations, Cmax and AUC0–180 min when D-allulose, D-sorbose and D-tagatose were orally administrated. KGA-2727, a selective SGLT1 inhibitor, did not affect the peripheral and portal vein concentrations and pharmacokinetic parameters of these rare sugars. The results suggest that D-allulose, D-sorbose and D-tagatose are likely transported via GLUT5 but not SGLT1 in rat small intestine. [ABSTRACT FROM AUTHOR]

Copyright of Modern Food Science & Technology is the property of Editorial Office of Modern Food Science & Technology and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

A one-pot extraction-assisted d-galactose-to-d-tagatose isomerization strategy was proposed based on the selective extraction of d-tagatose by phenylborate anions. 4-Vinylphenylboronic acid was selected with high extraction efficiency and selectivity towards d-tagatose. The extracted sugars could be desorbed through a two-staged stripping process with the purity of d-tagatose significantly increased. In-situ extraction-assisted d-galactose-to-d-tagatose isomerization was implemented for the first time ever reported, and the effect of boron-to-sugar ratio (boron: sugar) was investigated. The conversion yield of d-tagatose at 60 °C increased from ∼ 39 % (boron: sugar = 0.5) to ∼ 56 % (boron: sugar = 1) but then decreased to ∼ 44 % (boron: sugar = 1.5). With temperature increased to 70 °C, the conversion yield of d-tagatose was further improved to ∼ 61 % (boron: sugar = 1.5), with the minimized formation of byproducts. Moreover, high purity (∼83 %) and concentrated d-tagatose solution (∼40 g/L) was obtained after sequential desorption. The proposed extraction-assisted isomerization strategy achieved improving the yield and purity of d-tagatose, proving its feasibility in industrial applications.

Copyright of Journal of Dalian Polytechnic University is the property of Journal of Dalian Polytechnic University Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

L-Arabinose isomerase (L-AI) has been commonly used as an efficient biocatalyst to produce D-tagatose via the isomerization of D-galactose. However, it remains a significant challenge to efficiently synthesize D-tagatose using the native (wild type) L-AI at an industrial scale. Hence, it is extremely urgent to redesign L-AI to improve its catalytic efficiency towards D-galactose, and herein a structure-based molecular modification of Lactobacillus plantarum CY6 L-AI (LpAI) was performed. Among the engineered LpAI, both F118M and F279I mutants showed an increased D-galactose isomerization activity. Particularly, the specific activity of double mutant F118M/F279I towards D-galactose was increased by 210.1% compared to that of the wild type LpAI (WT). Besides the catalytic activity, the substrate preference of F118M/F279I was also largely changed from L-arabinose to D-galactose. In the enzymatic production of D-tagatose, the yield and conversion ratio of F118M/F279I were increased by 81.2% and 79.6%, respectively, compared to that of WT. Furthermore, the D-tagatose production of whole cells expressing F118M/F279I displayed about 2-fold higher than that of WT cell. These results revealed that the designed site-directed mutagenesis is useful for improving the catalytic efficiency of LpAI towards D-galactose.

To improve the performance of whey protein isolates (WPI) as an encapsulation agent in the food industry, we investigated the formation mechanism of WPI and D-tagatose (DT) conjugates using simultaneous rheological and Fourier-Transform infrared spectroscopy (FTIR) techniques, and analysed its effects in encapsulating quercetin (Que). The degree of glycosylation of the conjugates increased to 18 % after heating treatment, whereas particle size and potential decreased to 64 nm and −39 mV, with WPI/D10 showing the lowest value. Simultaneous rheological and Fourier transform infrared analyses showed that the structure of the WPI/DT complexes changed, and the gel properties were much better than those of WPI. Several new peaks appeared at 2961, 1525, 1450, 1392, and 1236 cm-1, indicating that adding DT affected the structure of WPI. DT promoted secondary structural changes in WPI by increasing the degree of hydrocarbon chain, O-H group vibration, and C-O stretching. The WPI/DT conjugates increased the solubility of Que to 60.74 % and ABTS clearance to 73.98 %. The study may offer a theoretical foundation for using WPI/DT-encapsulated Que in food industry. [Display omitted] • WPI/DT Maillard reaction product was used as the embedding material. • DT improved the structural, physical, and antioxidant properties of WPI/DT conjugate. • WPI/DT conjugates structure was tested by the simultaneous rheology-FTIR technology. • The technology indicated that the gel properties of WPI/DT conjugate were improved. • The conjugates improve the bio-accessibility and antioxidant properties of Que. [ABSTRACT FROM AUTHOR]

• Green tea extract EGCG potently prevents RIII in mouse model. • Oral EGCG reduces acute RIII severity in pelvic cancer patients undergoing RT. • The radio-protective effect of EGCG on intestine could be transferred via SFF transplantation. • EGCG enriches gut microbiota-derived metabolite D-tagatose. • D-tagatose prevents RIII via activation of AMPK signalling. Radiation-induced intestinal injury (RIII) compromises the clinical utility of pelvic radiotherapy (RT). We aimed to explore the protective effect and underlying mechanism of (−)-epigallocatechin-3-gallate (EGCG) on RIII. We evaluated the protective effect of EGCG on intestine in RIII mouse model and pelvic cancer patients, while explored the underlying mechanism through (1) 16S rRNA sequencing, (2) metabolomic profiles, (3) fresh sterile fecal filtrate (SFF) transplantation, and (4) transcriptome sequencing. EGCG efficiently prevented RIII in mouse, as reflected by improved survival, alleviated intestinal structure damage, promoted intestinal regeneration, and ameliorated gut microbiota dysbiosis. Prophylactic EGCG intervention reduced the severity of RIII in patients receiving pelvic RT. Mechanistically, the protective effect of EGCG could be transferred to other mice by SFF transplantation. EGCG enriched gut microbiota-derived metabolite D-tagatose, and oral administration of D-tagatose reproduced the radio-protective effect of EGCG via activating AMPK. Oral EGCG may be a promising strategy for preventing RIII clinically, and warrant further investigation in prospective randomized phase III trials. [ABSTRACT FROM AUTHOR]

Copyright of China Brewing is the property of China Brewing Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Chocolate is a popular and widely consumed product in different ages, however, the demand of consumers has increased for healthy chocolate with free or low sugar in recent years. The objective of present research is to substitute the sucrose of chocolate with D-tagatose and lemon essential oil for improving biological functions. The seven formulations of chocolate included distinct percentages of D-tagatose (0, 13, 29 and 47 %) and lemon essential oil (0, 0.1 and 0.2 %). Afterwards, the tests were performed on chocolate treatments such as moisture level, fat, sugar content before and after hydrolysis, color perception, hardness, antioxidant, microbial and sensory evaluations. The results demonstrated that moisture of treatments (except CTL5 containing 47.80 % D-tagatose and 0.1 % lemon essential oil) was noticeably different from the control. A significant difference was observed between the average sugars of treatments before hydrolysis, however, there was no considerable difference in fat levels (33.39 to 33.41 %). The essential oil has led to a reduction in hardness, so CTL6 (including 47.90 % D-tagatose and 0.2 % lemon essential oil) with higher essential oil had a lower hardness (16.61 N). D-tagatose and lemon essential oil had an influence on color features, but antioxidant functions were affected only by essential oil and all treatments were within the standard range in terms of microbial load. In sensory assessment, the overall acceptance score was given to the control (4.96) and CTL5 (4.92). The conclusion of present research introduced CTL5 as the most suitable treatment for production of healthy chocolate. [ABSTRACT FROM AUTHOR]

β-galactosidase is an enzyme capable of hydrolysing lactose, used in various branches of industry, mainly the food industry. As the efficient industrial use of enzymes depends on their reuse, it is necessary to find an effective method for immobilisation, maintaining high activity and stability. The present work proposes cross-linked magnetic cross-linked enzyme aggregates (mCLEAs) to prepare heterogeneous biocatalysts of β-galactosidase. Different concentrations of glutaraldehyde (0.6%, 1.0%, 1.5%), used as a cross-linking agent, were studied. The use of dextran-aldehyde as an alternative cross-linking agent was also evaluated. The mCLEAs presented increased recovered activity directly related to the concentration of glutaraldehyde. Modifications to the protocol to prepare mCLEAs with glutaraldehyde, adding a competitive inhibitor or polymer coating, have not been effective in increasing the recovered activity of the heterogeneous biocatalysts or its thermal stability. The biocatalyst prepared using dextran-aldehyde presented 73.6% recovered activity, aside from substrate affinity equivalent to the free enzyme. The thermal stability at 60 °C was higher for the biocatalyst prepared with glutaraldehyde (mCLEA-GLU-1.5) than the one produced with dextran-aldehyde (mCLEA-DEX), and the opposite happened at 50 °C. Results obtained for lactose hydrolysis, the use of its product to produce a rare sugar (D-tagatose) and operational and storage stability indicate that heterogeneous biocatalysts have adequate characteristics for industrial use. [ABSTRACT FROM AUTHOR]

d-Tagatose, a rare low-calorie sweetener, is ideal for beverages due to its high solubility and low viscosity. Current enzymatic production methods from d-galactose or d-galactitol are limited by reaction reversibility, affecting the yield and purity. This study demonstrates that Escherichia coli harbors a thermodynamically favorable pathway for producing d-tagatose from d-glucose via phosphorylation-epimerization-dephosphorylation steps. GatZ and KbaZ, annotated as aldolase chaperones, exhibit C4 epimerization activity, converting d-fructose-6-phosphate to d-tagatose-6-phosphate. Structural analysis reveals active site differences between these enzymes and class II aldolases, indicating functional divergence. By exploiting the strains' inability to metabolize d-tagatose, carbon starvation was applied to remove sugar byproducts. The engineered strains converted 45 g L -1 d-glucose to d-tagatose, achieving a titer of 7.3 g L -1 and a productivity of 0.1 g L -1 h -1 under test tube conditions. This approach highlights E. coli as a promising host for efficient d-tagatose production.

D-Tagatose is a representative rare sugar with the physiochemical properties of low energy and high sweetness, as well as excellent physiological functions such as blood sugar regulation, enhancement of intestinal flora, and prevention of dental caries. At present, D-tagatose production involves lactose hydrolysis and D-galactose isomerization processes, resulting in high production costs that hinder its industrial advancement. Tagatose 4-epimerase (T4Ease) has the capability to directly convert d-fructose into D-tagatose through C-4 epimerization, providing a new approach for D-tagatose production. In this study, a hyperthermostable T4Ease from Infirmifilum uzonense (Inuz-TE4ase) was identified from the Foldseek clustered AlphaFold database and its biochemical properties were characterized in detail. Under the optimal reaction conditions of 90 °C and pH 8.5 (Tris-HCl) with the addition of 1 mM Ni 2+ , the maximum catalytic activity towards d-fructose was determined to be 0.680 U/mg. Inuz-TE4ase exhibited exceptional thermostability, with half-life (t 1/2 ) values of 19.3 h at 85 °C and 8.9 h at 90 °C, respectively. Inuz-TE4ase was strictly metal-dependent, and its stability could be enhanced by Ni 2+ with an increase in the melting temperature (T m ) value from 101.1 °C to 105.7 °C. When 100 g/L d-fructose was used as the substrate, Inuz-TE4ase could catalyze the production of 21.67 g/L D-tagatose, indicating its significant potential for D-tagatose bioproduction., Competing Interests: Declaration of competing interest The authors declare no competing financial interest., (Copyright © 2025 Elsevier B.V. All rights reserved.)

d-Tagatose, a rare sugar endowed with a low-calorie property, superior taste quality, and probiotic functionality, has garnered significant research attention. However, the prevailing biological production methods relying on β-galactosidase and l-arabinose isomerase face challenges including high cost and suboptimal conversion efficiency. Consequently, it is of great research significance to find efficient alternative routes for d-tagatose synthesis. Previously, Thermotoga petrophila tagaturonate 3-epimerase was modified to function as tagatose 4-epimerase (T4E) enabling the direct conversion of d-fructose to d-tagatose. In this study, T4E was further engineered through directed evolution, specifically targeting the enhancement of its thermostability for application. This endeavor yielded promising T4E variants with superiority over those of the original enzyme. T4E I430P exhibits a half-life ( t 1/2 ) at 70 °C that is 1.83-fold that of T4E, with an increased melting temperature ( T m is 2.9 °C higher than T4E. Furthermore, whole-cell immobilization integrating these engineered T4E variants into a robust biocatalytic system was employed. This innovative approach not only underscores the practical feasibility of modifying enzymes through directed evolution but also establishes a foundation for the cost-effective, large-scale production of d-tagatose.t 1/2 was 1.69-fold that of T4E, and its T m is 2.9 °C higher than T4E. Furthermore, whole-cell immobilization integrating these engineered T4E variants into a robust biocatalytic system was employed. This innovative approach not only underscores the practical feasibility of modifying enzymes through directed evolution but also establishes a foundation for the cost-effective, large-scale production of d-tagatose.

Abstract As a rare low‐calorie sugar with special medicinal value, d‐tagatose is widely used in the field of food, beverages, medicine, and cosmetics. However, enzymatic d‐tagatose production in vitro is commonly limited to low conversion efficiency and poor thermo‐stability. Herein, taking advantage of the self‐assembling property of protein scaffold EutM (ethanolamine bacterial microcompartments), Spy and Snoop peptide pairs was used to drive the linkage between the EutM and cargo proteins, β‐galactosidase (BagB), and l‐arabinose isomerase (TMAI) to construct a dual‐enzymes cascade and realize the d‐tagatose production from lactose. The optimal conditions of the cascade were shown to be pH of 8.0, temperature of 60°C, 100 g/L lactose as substrate with supplementing 5 mM Mn2+. When the ratio of immobilized enzymes to EutM scaffold reached 1:6, the d‐tagatose productivity of the dual‐enzymes cascade could reach 1.03 g/L/h, which was 1.24‐fold higher than free enzymes. In addition, the EutM‐based scaffold could efficiently improve the stability of immobilized enzymes, in which 45% of the activity remained after 12 h, 2.14‐fold higher than the free one. Overall, an attractive EutM‐based self‐assembling platform immobilizing BagB and TMAI was developed, showing enhanced catalysis efficiency and enzyme thermo‐stability for d‐tagatose production.

Rare sugar was defined as a sugar that occurs in very small quantities in nature. Among them, l -ribose and d -tagatose were of high added value and useful as pharmaceutical intermediate for anti-HBV drugs or low calorie sweetener in food industry. Bio-production of the two rare sugar from biomass waste has not been investigated. Hence, development of a feasible and efficient co-production method was of practical usage. However, lack of suitable biocatalyst has become a bottleneck. By sequence alignment and analysis, a C-terminal α-helix from l -arabinose isomerase (L-AI) family was selected as a tool for protein engineering. This α-helix was ligated to C-terminal of Lactobacillus fermentum L-AI (LFAI) and significantly enhanced its thermostability and robustness for both l -arabinose and galactose catalysis. The mutant LFAI-C4 enzyme was immobilized by alginate and antimicrobial peptide poly- l -lysine, and was used to convert pretreated corncob acid hydrolysate (PCAH) into l -ribulose and d -tagatose in the presence of boric acid. In addition, we identified and immobilized a novel thermostable mannose-6-phosphate isomerase from Bacillus subtilis (BsMPI-2) which was efficient in catalyzing retaining l -ribulose into l -ribose and showing no activity on d -tagatose. The dual immobilized enzymes (LFAI-C4 and BsMPI-2) system co-produced 191.9 g/L l -ribose and 80.1 g/L d -tagatose, respectively. Showing a total yield of 46.6% from l -arabinose to l -ribose, which was the highest among reported. The dual immobilized enzymes system preserved 82% activity after 40 batches reaction, showing excellent potentials for industrial use. This study presents a promising alternative for rare sugar production from low-value raw material and showed satisfied conversion rate, product concentration, and operation stability. • An α-helix ligated to l -arabinose isomerase significantly enhanced thermostability and robustness. • The modified L-AI was used to convert pretreated corncob acid hydrolysate into l -ribulose and d -tagatose in high efficiency. • A novel thermostable mannose-6-phosphate isomerase from Bacillus subtilis was identified. • The dual L-AI and MPI immobilized enzymes system co-produced high concentrations of rare sugar. • The dual immobilized enzymes system preserved most activity after long term bio-reaction. [ABSTRACT FROM AUTHOR]

Diabetes mellitus is the most common metabolic disorder contributing to significant morbidity and mortality in humans. Different preventive and therapeutic agents, as well as various pharmacological strategies or non-pharmacological tools, improve the glycemic profile of diabetic patients. Isomaltulose, d-tagatose, and trehalose are naturally occurring, low glycemic sugars that are not synthesized by humans but widely used in food industries. Various studies have shown that these carbohydrates can regulate glucose metabolism and provide support in maintaining glucose homeostasis in patients with diabetes, but also can improve insulin response, subsequently leading to better control of hyperglycemia. In this review, we discussed the anti-hyperglycemic effects of isomaltulose, D-tagatose, and trehalose, comparing their properties with other known sweeteners, and highlighting their importance for the development of the pharmaceutical and food industries. [ABSTRACT FROM AUTHOR]

As a rare sugar, D-Tagatose is regarded as an excellent low-energy food sweetener. It is especially beneficial for physical health, such as inhibiting hyperglycemia, improving intestinal flora, and avoiding caries. It is broadly employed in food, medicine, cosmetics, and other fields. Acquiring D-Tagatose solubility is vital to develop D-Tagatose crystallization process to obtain high-quality D-Tagatose production. In this study, the solubility of D-Tagatose in seven pure solvents (methanol, ethanol, n-propanol, water, N, N-dimethylformamide, dimethyl sulfoxide, N-methylpyrrolidone), three binary solvents (ethanol-water mixtures, ethanol-methanol mixtures, and n-propanol-water mixtures) was determined by HPLC method at temperatures from 293.15 K to 323.15 K. Then, the Van't Hoff equation, Apelblat equation, λh equation, NRTL equation, CNIBS/Redlich-Kister model and modified Jouyban-Acree-van't Hoff model were used to correlate the solubility. The correlated solubility accounted for suitable compatibility with the experimental results (ARD

Acute respiratory distress syndrome (ARDS) is characterized by disruption of the alveolar-capillary barrier, resulting in severe alveolar edema and inflammation. D-tagatose (TAG) is a low-calorie fructose isomer with diverse biological activities whose role in ARDS has never been explored. We found that TAG protects lung tissues from injury in the oleic acid-induced rat model of ARDS. Seventeen male Sprague-Dawley rats were randomly assigned to 3 groups: Sham (n = 5), ARDS (n = 6), and TAG + ARDS (n = 6). The treatment groups were injected with oleic acid to induce ARDS, and the TAG + ARDS group was given TAG 3 days before the induction. After the treatments, the effect of TAG was evaluated by blood gas analysis and observing the gross and histological structure of the lung. The results showed that TAG significantly improved the oxygenation function, reduced the respiratory acidosis and the inflammatory response. TAG also improved the vascular permeability in ARDS rats and promoted the differentiation of alveolar type II cells, maintaining the stability of the alveolar structure. This protective effect of TAG on the lung may be achieved by activating the PTEN/PI3K/AKT pathway. Thus, TAG protects against oleic acid-induced ARDS in rats, suggesting a new clinical strategy for treating the condition. [ABSTRACT FROM AUTHOR]

The application of agarose oligosaccharides has garnered great attention, with their biological activities varying among different structures. However, it still meets a great bottleneck for the targeted production of odd-numbered neoagarooligosaccharides (NAOSs), such as neoagarotriose (NA3), due to the lack of one-step hydrolases. In this work, the α-agarase AgaA33 and β-galactosidase BgaD were synergistically used to prepare NA3 with agarose as a substrate. Additionally, an l-arabinose isomerase CaLAI from Clostridium acetobutylicum was characterized to valorize low-value byproducts (d-galactose) by forming d-tagatose, which exhibited good thermal stability without the need for additional metal ions. Under the optimal reaction conditions, the production of NA3 and d-galactose catalyzed by these three enzymes was 0.40 and 0.15 g/L, respectively. The artificial three-enzyme-based cascade transformation system not only achieved the highest production of NA3 until now but also allowed for the valorization of d-galactose, providing a wiser application route for agarose utilization.

Purpose: D-tagatose is an alternative sweetener with superior properties for reducing overall sugar absorption, which is especially important in the diabetic population. Large-scale production of D-tagatose remains costly, thereby limiting its commercialization. Introducing the unit operation of spray-drying may reduce some of these costs. However, D-tagatose is challenging to spray-dry due to its hydrophilic nature and low glass transition temperature (Tg). An appropriate spray-drying feed solution may offer an alternative approach if the material produced is glassy in nature, thus avoiding stickiness issues associated with Tgs less than the outlet temperature of the spray-dryer. Proof-of-concept formulations and processing conditions are presented in this study. Methods: D-tagatose was formulated with several excipients and screened for a sufficiently high Tg as characterized by differential scanning calorimetry (DSC). The spray-dry formulations were further characterized using attenuated total reflectance Fourier transform infrared spectrometry (ATR-FTIR), powder X-ray diffraction (PXRD), and scanning electron microscopy (SEM), and several viable options were identified. Results: D-tagatose formulated with HPMCAS and K90 both raised the Tg mix ( T g mix ) to within the processing range of spray-drying. These formulations were found to be sufficiently soluble in acetone/water cosolvent systems allowing for the production of processable powder. Specifically, SEM results confirm that both D-tagatose-HPMCAS and D-tagatose-K90 formulations result in particle formation using a small-scale dryer. Large-scale spray-drying is expected to allow for a greater processing range in comparison to small scale, suggesting that this approach will help commercialization. ATR-FTIR and PXRD results suggested that spray-dried D-tagatose-K90 composites lacked crystallinity, unlike the D-tagatose-HPMCAS composites. Conclusions: The addition of functional excipients allowed D-tagatose composites to be harvested using a typical laboratory-scale spray-dryer equipped with a bi-fluid nozzle. D-tagatose-K90 lacks crystallinity, while crystallinity is present in D-tagatose-HPMCAS composites. Crystallinity may not be a critical factor (depending on the future application) given the high aqueous solubility and high loading of the sugar. Both composites retained a sweet taste, suggesting that this spray-drying method may be worth further development. Other polymers with high Tg may be feasible as composites as well—as long as the amorphous phase or the residual amorphous phase has a Tg greater than, or about equal to, the outlet temperature of the spray-dryer. [ABSTRACT FROM AUTHOR]

• The D-tagaturonate epimerase (UxaE), which has a C4 epimerization function, catalyzes the synthesis of d -tagatose. • Semi-rational design has optimized UxaE to a 10.8-fold increase in efficiency. • Reshaping the substrate-binding pocket can effectively enhance the 4 epimerization function of the epimerase. d -tagatose, a rare sugar with broad applications in the food and pharmaceutical industries, can be synthesized from d -fructose by epimerases, an approach with significant biosynthesis potential. However, so far there has been only one attempt to engineer tagatose 4‑epimerase through random mutagenesis. The lack of exploration has severely restricted the application of epimerases in the industrial production of d -tagatose. The previous research primarily focused on enhancing the catalytic activity of epimerases through random mutagenesis, with limited understanding of how substrate-adjacent residues affect enzyme efficiency, which was hence the subject of this study. First, using a semi-rational design approach, after three rounds of mutagenesis and screening, the mutant (S125D/S268A/Q124E) of D-tagaturonate epimerase (UxaE) from Thermotoga neapolitana was obtained, which increased d -tagatose production efficiency by 10.8-fold compared to the wild type, with a specific activity of 1505.6 U/g, representing the highest specific activity reported for a UxaE mutant to date. Molecular dynamics simulations showed that the substitution of Ser to Asp-125 enhanced substrate-enzyme interactions, while the substitution of Glu to Gln-124 reinforced the hydrogen bond network around the catalytic residue Glu-128, further reshaping the substrate-binding pocket and enhancing catalytic activity. This study for the first time explored the enhancement of the C4 epimerization function of epimerases by reshaping their substrate-binding pockets, thereby providing a feasible route for the industrial biosynthesis of d -tagatose. [Display omitted] [ABSTRACT FROM AUTHOR]

d-Tagatose is a highly promising functional sweetener known for its various physiological functions. In this study, a novel tagatose 4-epimerase from Thermoprotei archaeon (Thar-T4Ease), with the ability to convert d-fructose to d-tagatose, was discovered through a combination of structure similarity search and sequence-based protein clustering. The recombinant Thar-T4Ease exhibited optimal activity at pH 8.5 and 85 °C, in the presence of 1 mM Ni 2+ . Its k cat and k cat / K m values toward d-fructose were measured to be 248.5 min -1 value of 198 h at 80 °C. Moreover, it achieved a conversion ratio of 18.9% using 100 g/L d-fructose as the substrate. Finally, based on sequence and structure analysis, crucial residues for the catalytic activity of Thar-T4Ease were identified by molecular docking and site-directed mutagenesis. This research expands the repertoire of enzymes with C4-epimerization activity and opens up new possibilities for the cost-effective production of d-tagatose from d-fructose.-1 ·min -1 , respectively. Notably, Thar-T4Ease exhibited remarkable thermostability, with a t 1/2 value of 198 h at 80 °C. Moreover, it achieved a conversion ratio of 18.9% using 100 g/L d-fructose as the substrate. Finally, based on sequence and structure analysis, crucial residues for the catalytic activity of Thar-T4Ease were identified by molecular docking and site-directed mutagenesis. This research expands the repertoire of enzymes with C4-epimerization activity and opens up new possibilities for the cost-effective production of d-tagatose from d-fructose.

d-Tagatose, a functional sweetener, is converted from d-galactose by l-arabinose isomerase, which catalyzes the conversion of l-arabinose to l-ribulose. In this study, the araA gene encoding l-arabinose isomerase from Klebsiella pneumoniae was cloned and expressed in Escherichia coli, and the expressed enzyme was purified and characterized. The purified l-arabinose isomerase, a soluble protein with 11.6-fold purification and a 22% final yield, displayed a specific activity of 1.8 U/mg for d-galactose and existed as a homohexamer of 336 kDa. The enzyme exhibited maximum activity at pH 8.0 and 40 °C in the presence of Mn2+ and relative activity for pentoses and hexoses in the order l-arabinose > d-galactose > l-ribulose > d-xylulose > d-xylose > d-tagatose > d-glucose. The thermal stability of recombinant E. coli cells expressing l-arabinose isomerase from K. pneumoniae was higher than that of the enzyme. Thus, the reaction conditions of the recombinant cells were optimized to pH 8.0, 50 °C, and 4 g/L cell concentration using 100 g/L d-galactose with 1 mM Mn2+. Under these conditions, 33.5 g/L d-tagatose was produced from d-galactose with 33.5% molar yield and 67 g/L/h productivity. Our findings will help produce d-tagatose using whole-cell reactions, extending its industrial application.

The one-pot production of tagatose from lactose using recombinant l -arabinose isomerase (L-AI) from Thermotoga maritima and commercial β-galactosidase (β-gal) from Aspergillus oryzae was evaluated. The β-gal and L-AI ratio (R β-gal/L-AI), initial lactose concentration and temperature were chosen as study variables. Maximum values of tagatose yield (Y Tag), specific tagatose productivity (π Tag) and sugars ratio (R Tag/Gal) obtained were 23 %, 0.202 ± 0.03 mmol Tag/(g prot⋅h) and 1, respectively. The composition of the product on a dry basis was as follows (g/g): 0.23 tagatose, 0.232 galactose, 0.454 glucose, 0.043 fructose, 0.029 lactose and 0.012 GOS. The one-pot production was superior compared with a sequential operation, where values of 16.5 %, 0.11 mmol Tag/(g prot⋅h) and 0.59 were obtained for Y Tag , π Tag and R Tag/Gal , respectively. • Tagatose was produced in one-pot operation with A. oryzae βgal and T. maritima L-AI. • Higher yield and specific productivity were achieved in one-pot operation. • Thermodynamically and kinetically controlled reaction are involved in one-pot operation. [ABSTRACT FROM AUTHOR]

A novel l -arabinose isomerase (L-AI) from Arthrobacter psychrolactophilus (Ap L-AI) was successfully cloned and characterized. The enzyme catalyzes the isomerization of d -galactose into a rare sugar d -tagatose. The recombinant Ap L-AI had an approximate molecular weight of about 258 kDa, suggesting it was an aggregate of five 58 kDa monomers and became the first record as a homo-pentamer L-AI. The catalytic efficiency (k cat /Km) and Km for d -galactose were 0.32 mM−1 min−1 and 51.43 mM, respectively, while for l -arabinose, were 0.64 mM−1 min−1 and 23.41 mM, respectively. It had the highest activity at pH 7.0–7.5 and 60 °C in the presence of 0.250 mM Mn2+. Ap L-AI was discovered to be an outstanding thermostable enzyme that only lost its half-life value at 60 °C for >1000 min. These findings suggest that l -arabinose isomerase from Arthrobacter psychrolactophilus is a promising candidate for d -tagatose mass-production due to its industrially competitive temperature. • Sequence of L-AI retrieved from the genome of Arthrobacter psychrolactophilus B7 • Gene cloning and overexpression in E. coli host using pET-28a(+) • Enzyme activity characterization indicated high thermal stability. [ABSTRACT FROM AUTHOR]

We designed and constructed a green and sustainable bioprocess to efficiently coproduce D -tagatose, bioethanol, and microbial protein from whey powder. First, a one-pot biosynthesis process involving lactose hydrolysis and D -galactose redox reactions for D -tagatose production was established in vitro via a three-enzyme cascade. Second, a nicotinamide adenine dinucleotide phosphate-dependent galactitol dehydrogenase mutant, D36A/I37R, based on the nicotinamide adenine dinucleotide-dependent polyol dehydrogenase from Paracoccus denitrificans was created through rational design and screening. Moreover, an NADPH recycling module was created in the oxidoreductive pathway, and the tagatose yield increased by 3.35-fold compared with that achieved through the pathway without the cofactor cycle. The reaction process was accelerated using an enzyme assembly with a glycine-serine linker, and the tagatose production rate was 9.28-fold higher than the initial yield. Finally, Saccharomyces cerevisiae was introduced into the reaction solution, and 266.5 g of D -tagatose, 162.6 g of bioethanol, and 215.4 g of dry yeast (including 38% protein) were obtained from 1 kg of whey powder (including 810 g lactose). This study provides a promising sustainable process for functional food ( D -tagatose) production. Moreover, this process fully utilized whey powder, demonstrating good atom economy., (© 2024 Wiley-VCH GmbH.)

[Display omitted] • A high-performance galactitol dehydrogenase was identified and charactered. • Redox pathway was constructed in Bacillus subtilis for D-tagatose production from D-galactose. • Fine-tuned promoter and cofactor regeneration provided 6.75-fold higher D-tagatose. • Optimum fermentation conditions and medium composition produced 39.57 g/L D-tagatose, providing a potential approach for D-tagatose production. D-tagatose holds significant importance as a functional monosaccharide with diverse applications in food, medicine, and other fields. This study aimed to explore the oxidoreductive pathway for D-tagatose production, surpassing the contemporary isomerization-mediated biosynthesis approach in order to enhance the thermodynamic equilibrium of the reactions. Initially, a novel galactitol dehydrogenase was discovered through biochemical and bioinformatics analyses. By co-expressing the galactitol dehydrogenase and xylose reductase, the oxidoreductive pathway for D-tagatose synthesis was successfully established in Bacillus subtilis. Subsequently, pathway fine-tuning was achieved via promoter regulation and dehydrogenase-mediated cofactor regeneration, resulting in 6.75-fold higher D-tagatose compared to that produced by the strain containing the unmodified promoter. Finally, optimization of fermentation conditions and medium composition produced 39.57 g/L D-tagatose in a fed-batch experiment, with a productivity of 0.33 g/L/h and a yield of 0.55 mol/mol D-galactose. These findings highlight the potential of the constructed redox pathway as an effective approach for D-tagatose production. [ABSTRACT FROM AUTHOR]

d -Tagatose is a novel low-calorie sweetener, but its application in milk and yogurt has not been explored. A novel l -arabinose isomerase gene was identified in Lactobacillus rhamnosus. The purified l -arabinose isomerase showed good thermal stability from 50 °C to 65 °C, and pH stability from 6.0 to 8.0. To prepare sweet milk, 3.77 × 10−7 kat/mL β-galactosidase and 9.02 × 10−8 kat/mL l -arabinose isomerase were added. After reaction at 65 °C for 3.5 h, the contents of glucose, d -galactose, and d -tagatose in milk were 25.00 g/L, 13.76 g/L, and 11.24 g/L, respectively, and lactose was not detected. Furthermore, Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus , with the respective characteristic of d -galactose, and glucose utilization were used as starters for sweet yogurt preparation. After fermentation, the yogurt contained 13.88 g/L of glucose, 10.13 g/L of d -galactose, and 10.90 g/L of d -tagatose, respectively. These results indicated the L. rhamnosus l -arabinose isomerase had a great potential for industrial application in sweetened milk and yogurt preparation. • A l -arabinose isomerase derived from L. rhamnosus was expressed in E. coli. • The enzyme had a high optimum temperature of 65 °C and good thermal stability. • Combined action of l -arabinose isomerase and β-galactosidase was explored. • The sweet milk and yogurt containing d -tagatose were prepared by the enzymes. [ABSTRACT FROM AUTHOR]

• Using DT for Maillard reaction with WPI to improve the efficiency of modified WPI. • Exploring the mechanism of WPI/DT gelation and thermal stability. • WPI/DT 3:1–1 has significantly improved gelability compared to WPI-1. • Improved thermal stability of all glycosylated samples. The Maillard reaction (MR) was used as a natural modification method to improve the functionality of proteins by covalent interaction with monosaccharides. This study aimed to prepare whey protein isolate (WPI)/ d -Tagatose (DT) conjugates by dry heating treatment to investigate the effect of different MR conditions on the structural changes of WPI and its effect on the WPI/DT conjugates gelation and thermal stability. Covalent interactions between WPI and DT were detected using multi-spectral methods. The MR conditions significantly affected the degree of glycosylation, structure, and molecular weight of the WPI/DT conjugates (P < 0.05). The MR changed the WPI/DT conjugates gel properties and thermal stability, where the WPI/DT 1:1–1 gel properties were stronger than those of WPI-1 and the heating deflection temperature after glycosylation was increased to 163.55 °C (P < 0.05). This study provides a theoretical basis for the application of DT as food ingredient in the development of WPI functional properties by MR. [ABSTRACT FROM AUTHOR]

• The effect of impurity sugars on the thermodynamics and kinetics of d -TAG crystallization has been studied. • Impurity sugars reduce the rate of d -TAG primary nucleation and crystal growth, but accelerate secondary nucleation rate. • The mechanism of the effect of impurity sugars on the nucleation and growth of d -TAG crystals was revealed by experiments and MD simulations. Impurity sugars produced in upstream process of functional sugars are significantly impacting the product quality. In this work, the effect of congeners (d -maltose (d -MAL), d -fructose (d -FRU), d -glucose (d -GLU)) on primary and secondary nucleation of d -tagatose (d -TAG) crystals was investigated. The impurity sugars showed an inhibition on primary nucleation of d -TAG crystals, while a promotion on the secondary nucleation of d -TAG. Interestingly, the impact of impurity sugars on d -TAG crystal growth was similar to that on primary nucleation. The diffusion ability, hydrogen bonding forming ability, interaction energy of d -TAG crystal surfaces and impurity sugars were evaluated by molecular dynamics (MD) simulations to reveal the nucleation and growth behavior. Based on the above findings, we designed the d -TAG crystallization experiments, and obtained d -TAG crystals with uniform particle size distribution and regular morphology. This study helps to understand the influence of impurity sugars on crystallization, guiding the industrial manufacturing of functional sugars. [ABSTRACT FROM AUTHOR]

L-arabinose isomerase (L-AI) is the key enzyme that isomerizes D-galactose to D-tagatose. In this study, to improve the activity of L-arabinose isomerase on D-galactose and its conversion rate in biotransformation, an L-arabinose isomerase from Lactobacillus fermentum CGMCC2921 was recombinantly expressed and applied in biotransformation. Moreover, its substrate binding pocket was rationally designed to improve the affinity and catalytic activity on D-galactose. We show that the conversion of D-galactose by variant F279I was increased 1.4 times that of the wild-type enzyme. The K m and k cat values of the double mutant M185A/F279I obtained by superimposed mutation were 530.8 mmol/L and 19.9 s -1 , respectively, and the catalytic efficiency was increased 8.2 times that of the wild type. When 400 g/L lactose was used as the substrate, the conversion rate of M185A/F279I reached a high level of 22.8%, which shows great application potential for the enzymatic production of tagatose from lactose.

d-Tagatose is one of the several healthy sweeteners that can be a substitute for sucrose and fructose in our daily life. Whole cell-catalyzed phosphorylation and dephosphorylation previously reported by our group afford a thermodynamic-driven strategy to achieve tagatose production directly from starch with high product yields. Nonetheless, the poor structural stability of cells and difficulty in biocatalyst recycling restrict its practical application. Herein, an efficient and stable semiartificial cell factory (SACF) was developed by constructing an organosilica network (OSN) artificial shell on the cells bearing five thermophilic enzymes to produce tagatose. The OSN artificial shell, the thickness of which can be regulated by changing the tetraethyl silicate concentration, exhibited tunable permeability and superior mechanical strength. In contrast with cells, SACFs showed a relative activity of 99.5% and an extended half-life from 33.3 to 57.8 h. Over 50% of initial activity was retained after 20 reuses. The SACFs can catalyze seven consecutive reactions with tagatose yields of over 40.7% in field applications.

The evolution of the bacterial phosphotransferase system (PTS) linked to glycolysis is dependent on the availability of naturally occurring sugars. Although bacteria exhibit sugar specificities based on carbon catabolite repression, the acquisition and evolvability of the cellular sugar preference under conditions that are suboptimal for growth (e.g., environments rich in a rare sugar) are poorly understood. Here, we generated Escherichia coli mutants via a retro-aldol reaction to obtain progeny that can utilize the rare sugar d-tagatose. We detected a minimal set of adaptive mutations in the d-fructose-specific PTS to render E. coli capable of d-tagatose utilization. These E. coli mutant strains lost the tight regulation of both the d-fructose and N -acetyl-galactosamine PTS following deletions in the binding site of the catabolite repressor/activator protein (Cra) upstream from the fruBKA operon and in the agaR gene, encoding the N -acetylgalactosamine (GalNAc) repressor, respectively. Acquired d-tagatose catabolic pathways then underwent fine-tuned adaptation via an additional mutation in 1-phosphofructose kinase to adjust metabolic fluxes. We determined the evolutionary trajectory at the molecular level, providing insights into the mechanism by which enteric bacteria evolved a substrate preference for the rare sugar d-tagatose. Furthermore, the engineered E. coli mutant strain could serve as an in vivo high-throughput screening platform for engineering non-phosphosugar isomerases to produce rare sugars. IMPORTANCE Microorganisms generate energy through glycolysis, which might have preceded a rapid burst of evolution, including the evolution of cellular respiration in the primordial biosphere. However, little is known about the evolvability of cellular sugar preferences. Here, we generated Escherichia coli mutants via a retro-aldol reaction to obtain progeny that can utilize the rare sugar d-tagatose. Consequently, we identified mutational hot spots and determined the evolutionary trajectory at the molecular level. This provided insights into the mechanism by which enteric bacteria evolved substrate preferences for various sugars, accounting for the widespread occurrence of these taxa. Furthermore, the adaptive laboratory evolution-induced cellular chassis could serve as an in vivo high-throughput screening platform for engineering tailor-made non-phosphorylated sugar isomerases to produce low-calorigenic rare sugars showing antidiabetic, antihyperglycemic, and antitumor activities.

Phosphatases catalyze the irreversible dephosphorylation of phosphate-containing compounds, and hence can be applied as the final enzymatic step for the synthesis of various biochemicals. However, the extensive substrate spectrums of phosphatases impose a great challenge for efficient biomanufacturing. Characterization of phosphatases is therefore of extreme importance. In this study, MmPase, a putative HAD phosphatase from Methanothermobacter marburgensis , was expressed, purified, and characterized. Recombinant MmPase was readily expressed in Escherichia coli , and required metal ions such as Mn2+ or Mg2+ to function. MmPase worked optimally at 50 °C, pH 6.5, and exhibited a half-life of 6.5 h under this condition. Among all substrates tested, MmPase established the highest dephosphorylation activity against D -tagatose 6-phosphate, and was relatively specific for this substrate than for D -glucose 1-phosphate, D -glucose 6-phosphate, and D -fructose 6-phosphate. Therefore, MmPase was integrated into an in vitro synthetic enzymatic biosystem for the one-pot production of D -tagatose from maltodextrin, and achieved a product yield of 37.6%. Our studies of MmPase provided a promising strategy for the economic and efficient production of D -tagatose in the future. • We examined the enzymatic properties of a putative phosphatase from M. marburgensis. • This enzyme was thermostable and relatively specific for D -tagatose 6-phosphate. • This enzyme was applied in an in vitro synthetic biosystem to produce D -tagatose. [ABSTRACT FROM AUTHOR]

[Display omitted] • Cheese whey powder (CWP) was used to produce three value-added rare sugars. • D-tagatose was produced by feeding CWP during whole cell biotransformation with E. coli. • D-arabitol and galactitol were produced from residual sugars by fermentation with yeast. • Finally, 68.35 g/L D-tagatose, 60.12 g/L D-arabitol and 28.26 g/L galactitol were produced. • The net titer of the three value-added products was 156.73 g/L. Dairy industry waste has been explored as a cheap and attractive raw material to produce various commercially important rare sugars. In this study, a lactose-rich dairy byproduct, namely cheese whey powder (CWP), was microbially converted into three low caloric sweeteners using whole-cell and fermentation technologies. Firstly, the simultaneous lactose hydrolysis and isomerization of lactose-derived D-galactose into D-tagatose was performed by an engineered Escherichia coli strain co-expressing β-galactosidase and L-arabinose isomerase, which eventually produced 68.35 g/L D-tagatose during sequential feeding of CWP. Subsequently, the mixed syrup containing lactose-derived D-glucose and residual D-galactose was subjected to fermentation by Metschnikowia pulcherrima E1, which produced 60.12 g/L D-arabitol and 28.26 g/L galactitol. The net titer of the three rare sugars was 156.73 g/L from 300 g/L lactose (equivalent to 428.57 g/L CWP), which was equivalent to 1.12 mol product/mol lactose and 52.24% conversion efficiency in terms of lactose. [ABSTRACT FROM AUTHOR]

A new strain of Lactococcus lactis producing d -tagatose was isolated and identified. Its l -arabinose isomerase coding gene was cloned and expressed in Escherichia coli BL21 (DE3). The optimal temperature and pH of the purified enzyme were 50 °C and 8.0. To produce d -tagatose from lactose, β- d -galactosidases from Lc. Lactis , Lactiplantibacillus plantarum , and Streptococcus thermophilus were further incorporated into E. coli by two strategies. These β- d -galactosidases were fused to l -arabinose isomerase coupled with a peptide linker (GGGGS) 3. Meanwhile, they were co-expressed with l -arabinose isomerase using pETDuet-1 vector. Among these recombinant strains, the cell co-expressing l -arabinose isomerase and S. thermophilus β- d -galactosidase showed maximal activity. SDS-PAGE results confirmed that exogenous enzymes had the maximum soluble expression level in this strain. At the optimal condition, the conversion rate of d -tagatose from 300 g/L lactose achieved to 42.4%, and the volumetric productivity reached 4.28 g/L/h at 15 h. This research established a highly efficient biotransformation system to produce d -tagatose from lactose. [Display omitted] • A new strain producing d -tagatose was isolated and identified as L. lactis X-18. • Different β- d -galactosidases were co-expressed with AraA to produce d -tagatose. • E. coli AI-d3 had the maximal production. • The conversion rate from lactose was 42.4%, and the productivity was 4.28 g/L/h. [ABSTRACT FROM AUTHOR]

Food manufacturers are under increasing pressure to limit the amount of free sugars in their products. Many have reformulated products to replace sucrose, glucose and fructose with alternative sweeteners, but some of these have been associated with additional health concerns. Rare sugars are ‘monosaccharides and their derivatives that hardly exist in nature’, and there is increasing evidence that they could have health benefits. This review aimed to scope the existing literature in order to identify the most commonly researched rare sugars, to ascertain their proposed health benefits, mechanisms of action and potential uses and to highlight knowledge gaps. A process of iterative database searching identified fifty-five relevant articles. The reported effects of rare sugars were noted, along with details of the research methodologies conducted. Our results indicated that the most common rare sugars investigated are d-psicose and d-tagatose, with the potential health benefits divided into three topics: glycaemic control, body composition and CVD. All the rare sugars investigated have the potential to suppress postprandial elevation of blood glucose and improve glycaemic control in both human and animal models. Some animal studies have suggested that certain rare sugars may also improve lipid profiles, alter the gut microbiome and reduce pro-inflammatory cytokine expression. The present review demonstrates that rare sugars could play a role in reducing the development of obesity, type 2 diabetes and/or CVD. However, understanding of the mechanisms by which rare sugars may exert their effects is limited, and their effectiveness when used in reformulated products is unknown.

The use of natural bio-based compounds becomes an eco-friendly strategy to control plant diseases. Rare sugars would be promising compounds as inducers of plant "sweet immunity". The present study aimed to investigate the induced resistance of grapevine leaves against Plasmopara viticola and Botrytis cinerea by a rare sugar-based product (IFP48) and its active ingredient D-tagatose (TAG), in order to elucidate molecular mechanism involved in defense-related metabolic regulations before and after pathogen challenge. Data showed that spraying leaves with IFP48 and TAG lead to a significant reduction of downy mildew, but not of gray mold disease. The induced protection against P. viticola relies on IFP48's and to a lesser extent TAG's ability to potentiate the activation of salicylic acid- and jasmonic acid/ethylene-responsive genes and stilbene phytoalexin accumulation. Most of defense responses remained upregulated in IFP48-treated plants after infection with P. viticola, but inconsistent following challenge with B. cinerea. The beneficial effects of IFP48 were associated with an enhanced accumulation of tagatose inside leaf tissues compared to TAG treatment. Meanwhile, the amounts of sugars, glucose, fructose, maltose, galactose and trehalose remained unchanged or decreased in IFP48-treated leaves after P. viticola infection, although only a few genes involved in sugar transport and metabolism showed transcriptional regulation. This suggests a contribution of sugar homeostasis to the IFP48-induced sweet immune response and priming plants for enhanced resistance to P. viticola, but not to B. cinerea. [ABSTRACT FROM AUTHOR]

Objective: To examine the effects of d-tagatose or stevia preloads on carbohydrate metabolism markers after an oral glucose load, as well as subjective and objective appetite in women with insulin resistance (IR)., Research Design and Methods: Randomized controlled crossover study. Women with IR without T2DM (n = 33; aged 23.4 ± 3.8; BMI 28.1 ± 3.4 kg × m -2 ) underwent three oral glucose loads (3 h each) on three different days. Ten min before oral glucose load, volunteers consumed a preload of 60 mL water (control), 60 mL water with stevia (15.3 mg), or d-tagatose (5000 mg). Serum glucose and C-peptide were evaluated at -10, 30-, 60-, 90-, 120-, and 180-min. Subjective appetite was determined with a visual analog scale. Food intake was measured at ad libitum buffet after 180 min., Results: C-peptide iAUC was significantly higher for stevia (median (IQR): 1033 (711-1293) ng × min × L -1 ) vs. d-tagatose (794 (366-1134) ng × min × L -1 ; P = 0.001) or control (730 (516-1078) ng × min × L -1 ; P = 0.012). At 30- and 60-min serum glucose was higher for stevia vs other conditions (P < 0.01). Volunteers reported greater satiety for stevia and d-tagatose vs. control at 60 min and greater desire to eat for stevia vs. control at 120- min (all P < 0.05). Objective appetite did not vary by condition (P = 0.06)., Conclusions: Our findings suggest that these NNS are not inert. Stevia intake produced an acute response on C-peptide release while increased serum glucose at earlier times. It is possible that NNS affects subjective but not objective appetite. This trial is registered at clinicaltrials.gov as NCT04327245., Clinical Trial Registry: NCT04327245., Competing Interests: Declaration of competing interest The authors have no conflicts of interest to declare., (Copyright © 2022 European Society for Clinical Nutrition and Metabolism. Published by Elsevier Ltd. All rights reserved.)