RESUMO
Lytic polysaccharide monooxygenases (LPMOs) are copper-dependent enzymes that oxidatively cleave recalcitrant polysaccharides such as cellulose. Several studies have reported LPMO action in synergy with other carbohydrate-active enzymes (CAZymes) for the degradation of lignocellulosic biomass but direct LPMO action at the plant tissue level remains challenging to investigate. Here, we have developed a MALDI-MS imaging workflow to detect oxidised oligosaccharides released by a cellulose-active LPMO at cellular level on maize tissues. Using this workflow, we imaged LPMO action and gained insight into the spatial variation and relative abundance of oxidised and non-oxidised oligosaccharides. We reveal a targeted action of the LPMO related to the composition and organisation of plant cell walls.
Assuntos
Oxigenases de Função Mista , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Zea mays , Zea mays/química , Oxigenases de Função Mista/metabolismo , Oxigenases de Função Mista/química , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Celulose/química , Celulose/metabolismo , Parede Celular/química , Parede Celular/metabolismo , Oligossacarídeos/química , Oligossacarídeos/metabolismo , Lignina/química , Lignina/metabolismo , Oxirredução , Polissacarídeos/química , Polissacarídeos/metabolismo , Proteínas de Plantas/química , Proteínas de Plantas/metabolismoRESUMO
Alginate is one of the most important marine colloidal polysaccharides, and its oligosaccharides have been proven to possess diverse biological functions. Alginate lyases could specifically degrade alginate and therefore serve as desirable tools for the research and development of alginate. In this report, a novel catalytic domain, which demonstrated no significant sequence similarity with all previously defined functional domains, was verified to exhibit a random endo-acting lyase activity to alginate. The action pattern analysis revealed that the heterologously expressed protein, named Aly44A, preferred to degrade polyM. Its minimum substrates and the minimum products were identified as unsaturated alginate trisaccharides and disaccharides, respectively. Based on the sequence novelty of Aly44A and its homologs, a new polysaccharide lyase family (PL44) was proposed. The discovery of the novel enzyme and polysaccharide lyase family provided a new entrance for the gene-mining and acquiring of alginate lyases, and would facilitate to the utilization of alginate and its oligosaccharides.
Assuntos
Alginatos , Polissacarídeo-Liases , Polissacarídeo-Liases/metabolismo , Polissacarídeo-Liases/química , Polissacarídeo-Liases/genética , Alginatos/química , Alginatos/metabolismo , Especificidade por Substrato , Domínio Catalítico , Oligossacarídeos/química , Oligossacarídeos/metabolismo , Sequência de Aminoácidos , Ácidos Hexurônicos/química , Ácidos Hexurônicos/metabolismoRESUMO
Oligosaccharides from uronic acid-containing polysaccharides can be produced either by chemical or enzymatic degradation. The benefit of using enzymes, called lyases, is their high specificity for various glycosidic linkages. Lyases cleave the polysaccharide chain by an ß-elimination reaction, yielding oligosaccharides with an unsaturated sugar (4-deoxy-l-erythro-hex-4-enepyranosyluronate) at the non-reducing end. In this work we have systematically studied acid degradation of unsaturated uronic acid oligosaccharides. Based on these findings, a method for preparing saturated oligosaccharides by enzymatic degradation of uronic acid-containing polysaccharides was developed. This results in oligosaccharides with a pre-defined distribution and proportion of sugar residues compared to the products of chemical degradation, while maintaining the chemical structure of the non-reducing end. The described method was demonstrated for generating saturated oligosaccharides of alginate, heparin and polygalacturonic acid. In the case of alginate, the ratio of hydrolysis rate of Δ-G and Δ-M linkages to that of G-G and M-M linkages, respectively, was found to be approximately 65 and 43, at pH* 3.4, 90 °C. Finally, this method has been demonstrated to be superior in the production of α-l-guluronate oligosaccharides with a lower content of ß-d-mannuronate residues compared to what can be achieved using chemical depolymerization alone.
Assuntos
Alginatos , Oligossacarídeos , Ácidos Urônicos , Alginatos/química , Oligossacarídeos/química , Ácidos Urônicos/química , Hidrólise , Polissacarídeo-Liases/química , Polissacarídeo-Liases/metabolismo , Polissacarídeos/química , Pectinas/química , Heparina/químicaRESUMO
Non-digestible oligosaccharides (OS) and allulose have beneficial health properties and could reduce the amount of added sugar in baked goods. In this study allulose and various OS [fructo-oligosaccharides (FOS), galacto-oligosaccharides (GOS), lactosucrose (LOS), isomalto-oligosaccharides (IMO), Promitor 70R (P70R), and xylo-oligosaccharides (XOS)] were added to a wire-cut cookie formulation at concentrations determined to have similar effects on the gelatinization temperature (Tgel) of starch relative to sucrose. Different baking performance attributes of the doughs and cookies were assessed, including: appearance, spread, color, texture, and % moisture loss after baking. The results were correlated to: OS solution and solid properties and OS effects on starch thermal events (gelatinization, pasting, and retrogradation). The Tgel-matching formulation protocol was effective in producing reduced-sugar cookies which had similar appearance, color, and spread attributes compared to the sucrose control; however, cookie texture significantly varied. Cookies containing allulose were the least similar to the control, having darker color, reduced spread, and softer cake-like texture. The only OS cookies that matched the texture of the sucrose control contained LOS, while P70R cookies were the hardest. Cookie texture correlated strongly with the % total moisture loss after baking (r = -0.8763) and was best explained by OS solution viscosity: more viscous OS solutions limited moisture release and resulted in harder cookies. The Tgel of starch also correlated with OS solution viscosity (r = 0.7861) and should be accounted for in reduced sugar applications. The OS recommended as sucrose replacers in cookies based on principal component analysis groupings were: XOS > IMO > LOS > and GOS.
Assuntos
Oligossacarídeos , Oligossacarídeos/química , Culinária/métodos , Sacarose/química , Amido/química , Cor , Água/química , Frutose/química , Manipulação de Alimentos/métodos , Viscosidade , TemperaturaRESUMO
Human milk oligosaccharides (HMOs) are an evolutionarily significant advantage bestowed by mothers for facilitating the development of the infant's gut microbiota. They can avoid absorption in the stomach and small intestine, reaching the colon successfully, where they engage in close interactions with gut microbes. This process also enables HMOs to exert additional prebiotic effects, including regulating the mucus layer, promoting physical growth and brain development, as well as preventing and mitigating conditions such as NEC, allergies, and diarrhea. Here, we comprehensively review the primary ways by which gut microbiota, including Bifidobacteria and other genera, utilize HMOs, and we classify them into five central pathways. Furthermore, we emphasize the metabolic benefits of bacteria consuming HMOs, particularly the recently identified intrinsic link between HMOs and the metabolic conversion of tryptophan to indole and its derivatives. We also examine the extensive probiotic roles of HMOs and their recent research advancements, specifically concentrating on the unsummarized role of HMOs in regulating the mucus layer, where their interaction with the gut microbiota becomes crucial. Additionally, we delve into the principal tools used for functional mining of new HMOs. In conclusion, our study presents a thorough analysis of the interaction mechanism between HMOs and gut microbiota, emphasizing the cooperative utilization of HMOs by gut microbiota, and provides an overview of the subsequent probiotic effects of this interaction. This review provides new insights into the interaction of HMOs with the gut microbiota, which will inform the mechanisms by which HMOs function.
Assuntos
Microbioma Gastrointestinal , Leite Humano , Oligossacarídeos , Prebióticos , Humanos , Microbioma Gastrointestinal/fisiologia , Leite Humano/química , Leite Humano/microbiologia , Oligossacarídeos/química , Probióticos , Lactente , Bactérias/metabolismo , Bifidobacterium/fisiologiaRESUMO
The plant cell wall is rich in polysaccharides with high heterogeneity. Investigating the composition and structure of cell wall polysaccharides is crucial for understanding the functionalities of plant cell walls. Carbohydrate electrophoresis is a sensitive and rapid method to analyze polysaccharides qualitatively and quantitatively. The process includes digesting the polysaccharides with appropriate cleavage enzymes, labeling the reducing ends of the released oligosaccharides with a highly charged fluorophore, and separating the labeled oligosaccharides in a polyacrylamide gel via high-voltage electrophoresis. The generated fluorescence can be calculated as compared to that of oligosaccharide standards. Therefore, this is a convenient method for polysaccharide characterization that can be performed in most laboratories. Here, we introduce the detailed operational steps and precautions, which are helpful for researchers to quickly obtain the structural information of polysaccharides.
Assuntos
Parede Celular , Polissacarídeos , Parede Celular/química , Polissacarídeos/análise , Polissacarídeos/química , Oligossacarídeos/análise , Oligossacarídeos/química , Eletroforese em Gel de Poliacrilamida/métodos , Eletroforese/métodosRESUMO
Various Xanthomonas species cause well-known plant diseases. Among various pathogenic factors, the role of α-1,6-cyclized ß-1,2-glucohexadecaose (CßG16α) produced by Xanthomonas campestris pv. campestris was previously shown to be vital for infecting model organisms, Arabidopsis thaliana and Nicotiana benthamiana. However, enzymes responsible for biosynthesizing CßG16α are essentially unknown, which limits the generation of agrichemicals that inhibit CßG16α synthesis. In this study, we discovered that OpgD from X. campestris pv. campestris converts linear ß-1,2-glucan to CßG16α. Structural and functional analyses revealed OpgD from X. campestris pv. campestris possesses an anomer-inverting transglycosylation mechanism, which is unprecedented among glycoside hydrolase family enzymes.
Assuntos
Xanthomonas campestris , Xanthomonas campestris/enzimologia , Xanthomonas/enzimologia , Doenças das Plantas/microbiologia , Oligossacarídeos/química , Oligossacarídeos/metabolismo , Modelos MolecularesRESUMO
The hexasaccharide arabinan domain of Mycobacterial Arabinogalactan was provided with the versatile methodology toward ß-selective arabinofuranosylation directed by B(C6F5)3, demonstrating the effectiveness of the ß-arabinofuranosylation strategy. Derivatization of the amino moiety at the reducing end are essential prerequisites for elucidating the biosynthetic pathway and conjugating of this compound to a protein carrier for vaccine generation.
Assuntos
Galactanos , Galactanos/química , Galactanos/síntese química , Oligossacarídeos/síntese química , Oligossacarídeos/química , Sequência de Carboidratos , Mycobacterium/química , PolissacarídeosRESUMO
To effectively solve the problem of significant loss of transplanted cells caused by thrombosis during cell transplantation, this study simulates the human fibrinolytic system and combines metabolic oligosaccharide engineering with strain-promoted azide-alkyne cycloaddition (SPAAC) click chemistry to construct a cell surface with fibrinolytic activity. First, a copolymer (POL) of oligoethylene glycol methacrylate (OEGMA) and 6-amino-2-(2-methylamido)hexanoic acid (Lys) was synthesized by reversible addition-fragmentation chain transfer (RAFT) copolymerization, and the dibenzocyclooctyne (DBCO) functional group was introduced into the side chain of the copolymer through an active ester reaction, resulting in a functionalized copolymer DBCO-PEG4-POL with ε-lysine ligands. Then, azide functional groups were introduced onto the surface of HeLa model cells through metabolic oligosaccharide engineering, and DBCO-PEG4-POL was further specifically modified onto the surface of HeLa cells via the SPAAC "click" reaction. In vitro investigations revealed that compared with unmodified HeLa cells, modified cells not only resist the adsorption of nonspecific proteins such as fibrinogen and human serum albumin but also selectively bind to plasminogen in plasma while maintaining good cell viability and proliferative activity. More importantly, upon the activation of adsorbed plasminogen into plasmin, the modified cells exhibited remarkable fibrinolytic activity and were capable of promptly dissolving the primary thrombus formed on their surfaces. This research not only provides a novel approach for constructing transplantable cells with fibrinolytic activity but also offers a new perspective for effectively addressing the significant loss of transplanted cells caused by thrombosis.
Assuntos
Química Click , Reação de Cicloadição , Fibrinólise , Oligossacarídeos , Humanos , Células HeLa , Oligossacarídeos/química , Fibrinólise/efeitos dos fármacos , Engenharia Metabólica , Azidas/química , Polietilenoglicóis/química , Metacrilatos/química , Alcinos/química , Animais , Sobrevivência Celular/efeitos dos fármacos , Plasminogênio/química , Plasminogênio/metabolismo , Propriedades de SuperfícieRESUMO
We herein report for the first time the inter- and intramolecular orthogonal cleavage of two ortho-nitrobenzyl (NB) analogues. It is shown that the nitroveratryl (NV) group can be photolyzed with high priority when NV and ortho-nitrobenzyl carbonate (oNBC) are used together as the protecting groups of glycans. Notably, the photolytic products could be used directly in the subsequent glycosylation without further purification. With the above-mentioned orthogonal photolabile protecting group strategy in hand, a Mycobacterium tuberculosis tetrasaccharide and a derivative of glucosyl glycerol were rapidly prepared.
Assuntos
Mycobacterium tuberculosis , Oligossacarídeos , Glicosilação , Estrutura Molecular , Mycobacterium tuberculosis/química , Nitrobenzenos/química , Oligossacarídeos/química , Oligossacarídeos/síntese químicaAssuntos
Antígenos de Grupos Sanguíneos , Leite Humano , Norovirus , Humanos , Norovirus/efeitos dos fármacos , Norovirus/metabolismo , Leite Humano/química , Antígenos de Grupos Sanguíneos/metabolismo , Sítios de Ligação , Oligossacarídeos/metabolismo , Oligossacarídeos/química , Trissacarídeos/metabolismoRESUMO
Chemotherapy-induced mucositis (CIM) is the typical side effect of chemotherapy. This study investigates the potential of alginate oligosaccharide (AOS) in ameliorating CIM induced by 5-fluorouracil (5-FU) in a murine model and its underlying mechanisms. AOS effectively mitigated body weight loss and histopathological damage, modulated inflammatory cytokines and attenuated the oxidative stress. AOS restored intestinal barrier integrity through enhancing expression of tight junction proteins via MLCK signaling pathway. AOS alleviated intestinal mucosal damage by inhibiting TLR4/MyD88/NF-κB signaling pathway, downregulating the pro-apoptotic protein Bax and upregulating the anti-apoptotic protein Bcl-2. Moreover, AOS significantly enriched intestinal Akkermansiaceae and increased the production of short-chain fatty acids (SCFAs), most notably butyrate and isovalerate. Pre-treatment with butyrate and isovalerate also alleviated 5-FU-induced CIM. In conclusion, AOS effectively mitigated CIM through strenghthening intestinal barrier, attenuating inflammation, and modulating gut microbiota and intestianl levels of butyrate and isovalerate. These finding indicate that AOS could be potentially utilized as a supplemental strategy for prevention or mitigation of CIM.
Assuntos
Alginatos , Butiratos , Fluoruracila , Mucosa Intestinal , Mucosite , Oligossacarídeos , Fluoruracila/efeitos adversos , Animais , Mucosite/induzido quimicamente , Mucosite/tratamento farmacológico , Mucosite/metabolismo , Mucosite/patologia , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Camundongos , Oligossacarídeos/farmacologia , Oligossacarídeos/química , Butiratos/farmacologia , Butiratos/metabolismo , Alginatos/farmacologia , Alginatos/química , Microbioma Gastrointestinal/efeitos dos fármacos , Masculino , Estresse Oxidativo/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Citocinas/metabolismoRESUMO
In this study, ß-1,3-xylanase (Xyl3088) was designed and prepared by constructing the expression vector plasmid and expressing and purifying the fusion protein. ß-1,3-xylo-oligosaccharides were obtained through the specific enzymatic degradation of ß-1, 3-xylan from Caulerpa lentillifera. The enzymolysis conditions were established and optimized as follows: Tris-HCl solution 0.05â¯mol/L, temperature of 37⯰C, enzyme amount of 250⯵L, and enzymolysis time of 24â¯h. The oligosaccharides' compositions and structural characterization were identified by thin-layer chromatography (TLC), ion chromatography (IC) and liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS). The IC50 values for scavenging 1,1-diphenyl-2-picrylhydrazyl (DPPH), 2,2-azino-bis-3-ethyl-benzothiazoline-p-sulfonic acid (ABTS+), and superoxide anion radical (â¢O2-) were 13.108, 1.258, and 65.926â¯mg/mL for ß-1,3-xylo-oligosaccharides, respectively, and 27.588, 373.048, and 269.12â¯mg/mL for ß-1,4-xylo-oligosaccharides, respectively. Compared with ß-1,4-xylo-oligosaccharides, ß-1,3-xylo-oligosaccharides had substantial antioxidant activity and their antioxidant effects were concentration dependent. ß-1,3-xylo-oligosaccharides also possessed a stronger anti-inflammatory effect on RAW 264.7 cells stimulated by lipopolysaccharide (LPS) than ß-1,4-xylo-oligosaccharides. At a working concentration of 100⯵g/mL, ß-1,3-xylo-oligosaccharides inhibited the release of NO and affected the expression of IL-1ß, TNF-α, and other proteins secreted by cells, effectively promoting the release of pro-inflammatory mediators by immune cells in response to external stimuli and achieving anti-inflammatory effects. Therefore, ß-1,3-xylo-oligosaccharides are valuable products in food and pharmaceutical industries.
Assuntos
Oligossacarídeos , Camundongos , Oligossacarídeos/farmacologia , Oligossacarídeos/química , Animais , Células RAW 264.7 , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/química , Antioxidantes/farmacologia , Antioxidantes/química , Xilosidases/metabolismo , Xilosidases/genética , Xilosidases/química , Algas Comestíveis , CaulerpaRESUMO
Ovarian cancer, the deadliest gynecological malignancy, primarily treated with chemotherapy. However, systemic chemotherapy often leads to severe toxic side effects and chemoresistance. Drug-loaded aerogels have emerged as a promising method for drug delivery, as they can improve drug solubility and bioavailability, control drug release, and reduce drug distribution in non-targeted tissues, thereby minimizing side effects. In this research, chitosan oligosaccharide (COS)-loaded nanofibers composite chitosan (CS) aerogels (COS-NFs/CS) with a porous network structure were created using nanofiber recombination and freeze-drying techniques. The core layer of the aerogel has a COS loading rate of 60 %, enabling the COS-NFs/CS aerogel to significantly inhibit the migration and proliferation of ovarian cancer cells (resulting in a decrease in the survival rate of ovarian cancer cells to 33.70 % after 48 h). The coaxial fiber's unique shell-core structure and the aerogel's porous network structure enable the COS-NFs/CS aerogels to release COS steadily and slowly over 30 days, effectively reducing the initial burst release of COS. Additionally, the COS-NFs/CS aerogels exhibit good biocompatibility, degradability (only retaining 18.52 % of their weight after 6 weeks of implantation), and promote angiogenesis, thus promoting wound healing post-oophorectomy. In conclusion, COS-NFs/CS aerogels show great potential for application in the treatment of ovarian cancer.
Assuntos
Quitosana , Preparações de Ação Retardada , Nanofibras , Oligossacarídeos , Neoplasias Ovarianas , Quitosana/química , Feminino , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/patologia , Porosidade , Nanofibras/química , Humanos , Oligossacarídeos/química , Animais , Preparações de Ação Retardada/química , Preparações de Ação Retardada/farmacologia , Liberação Controlada de Fármacos , Linhagem Celular Tumoral , Géis/química , Movimento Celular/efeitos dos fármacos , Antineoplásicos/farmacologia , Antineoplásicos/química , Portadores de Fármacos/química , Camundongos , Proliferação de Células/efeitos dos fármacosRESUMO
Designing thermo-responsive nanocarriers based on biopolymers is fascinating and challenging for cancer therapy. In this study, thermo-responsive composite nanoparticles (CNPs) were prepared using hydroxybutyl chitosan oligosaccharide (HBCOS) and sodium caseinate (SC) via electrostatic interactions and covalent crosslinking. The temperature-responsive behaviors of CNPs were induced by the breakage of hydrogen bonds and the shrinkage of chains in nanoparticles. The CNPs exhibited concentration-independent thermo-responsive behavior, non-adsorption aggregation, and non-hemolysis, suggesting excellent stability and thermo-sensitivity. The initial release rate and final amount of DOX released from CNPs at 42 °C were higher than that at 37 °C, showing a thermo-responsive release, which was also more prominent at lower pH. The release of DOX from CNPs followed first order kinetics based on Fickian diffusion. In vitro cytotoxicity assays confirmed the thermo-responsive antitumor activity of DOX-loaded CNPs as the HT-29 cell viability incubated with DOX-loaded CNPs at 42 °C was significantly lower than that at 37 °C. Cellular uptake experiments proved that DOX-loaded CNPs accumulated in the cytoplasm after being endocytosed and promoted DOX release by increasing environment temperature. This study generated stable thermo-sensitive CNPs based on biopolymers, which can be used as potential nanocarriers for the controlled release of anticancer drugs for cancer therapy.
Assuntos
Quitosana , Doxorrubicina , Liberação Controlada de Fármacos , Nanopartículas , Oligossacarídeos , Temperatura , Quitosana/química , Humanos , Nanopartículas/química , Doxorrubicina/química , Doxorrubicina/farmacologia , Oligossacarídeos/química , Oligossacarídeos/farmacologia , Portadores de Fármacos/química , Antineoplásicos/farmacologia , Antineoplásicos/química , Células HT29 , Neoplasias/tratamento farmacológico , Concentração de Íons de Hidrogênio , Sobrevivência Celular/efeitos dos fármacosRESUMO
In the present study, different oligosaccharides (fructooligosaccharide (FOS), galactooligosaccharide (GOS), isomaltooligosaccharide (IMO), and xylooligosaccharide (XOS)) were modified on casein (CN) via Maillard reaction. The CN-oligosaccharide conjugates were evaluated for modifications to functional groups, fluorescence intensity, water- and oil-holding properties, emulsion foaming properties, as well as general emulsion properties and stability. The results demonstrated that the covalent combination of CN and oligosaccharides augmented the spatial repulsion and altered the hydrophobic milieu of proteins, which resulted in a diminution in water-holding capacity, an augmentation in oil-holding capacity, and an enhancement in the emulsification properties of proteins. Among them, CN-XOS exhibited the most pronounced changes, with the emulsification activity index and emulsion stability index increasing by approximately 72% and 84.3%, respectively. Furthermore, CN-XOS emulsions have smaller droplet sizes and higher absolute potential values than CN emulsions. Additionally, CN-XOS emulsions demonstrate remarkable stability when ion concentration and pH are varied. These findings indicate that oligosaccharides modified via Maillard reaction can be used as good natural emulsifiers. This provides a theoretical basis for using oligosaccharides to modify proteins and act as natural emulsifiers.
Assuntos
Caseínas , Emulsificantes , Emulsões , Reação de Maillard , Oligossacarídeos , Oligossacarídeos/química , Caseínas/química , Emulsificantes/química , Emulsões/química , Concentração de Íons de Hidrogênio , Tamanho da Partícula , Interações Hidrofóbicas e Hidrofílicas , Água/químicaRESUMO
Soaking pulses in water is a traditional practice widely used both by many households and by the food industry, and depending on the specific conditions used, can effectively reduce α-galactosides. Monitoring changes in α-galactoside content in pulses under different steeping conditions can provide insights into the degradation mechanisms and help overcome the barrier to consumption caused by digestive problems. In this study, we analyzed the impact of steeping at different temperatures (30, 45, 60, 75, and 90 °C) and at different pH (4.0, 5.0, and 6.0) on α-galactosides content in chickpeas, lentils, and beans. Our results showed that the lower the pH, the faster the α-galactosides were reduced. Moreover, steeping at lower temperatures (30 °C and 45 °C) favored hydrolysis of α-galactosides, whereas steeping at higher temperatures (60, 75, and 90 °C) favored diffusion. Soaking at 45 °C at a pH of 4.0 for 3 h resulted in acceptable levels of α-galactosides (less than 1 g/100 g), i.e. a reduction of up to 65 % in chickpeas, 85 % in lentils, and 52 % in beans.
Assuntos
Cicer , Lens (Planta) , Oligossacarídeos , Rafinose , Temperatura , Concentração de Íons de Hidrogênio , Hidrólise , Rafinose/química , Rafinose/análise , Oligossacarídeos/química , Oligossacarídeos/análise , Lens (Planta)/química , Cicer/química , Manipulação de Alimentos/métodos , Galactosídeos/química , DifusãoRESUMO
Here we describe a complex enzymatic approach to the efficient transformation of abundant waste chitin, a byproduct of the food industry, into valuable chitooligomers with a degree of polymerization (DP) ranging from 6 to 11. This method involves a three-step process: initial hydrolysis of chitin using engineered variants of a novel fungal chitinase from Talaromyces flavus to generate low-DP chitooligomers, followed by an extension to the desired DP using the high-yielding Y445N variant of ß-N-acetylhexosaminidase from Aspergillus oryzae, achieving yields of up to 57%. Subsequently, enzymatic deacetylation of chitooligomers with DP 6 and 7 was accomplished using peptidoglycan deacetylase from Bacillus subtilis BsPdaC. The innovative enzymatic procedure demonstrates a sustainable and feasible route for converting waste chitin into unavailable bioactive chitooligomers potentially applicable as natural pesticides in ecological and sustainable agriculture.
Assuntos
Aspergillus oryzae , Quitina , Quitinases , Proteínas Fúngicas , Oligossacarídeos , Talaromyces , Quitina/metabolismo , Quitina/química , Quitinases/metabolismo , Quitinases/genética , Quitinases/química , Talaromyces/enzimologia , Talaromyces/genética , Talaromyces/química , Talaromyces/metabolismo , Oligossacarídeos/metabolismo , Oligossacarídeos/química , Hidrólise , Aspergillus oryzae/enzimologia , Aspergillus oryzae/genética , Aspergillus oryzae/metabolismo , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/genética , Proteínas Fúngicas/química , Bacillus subtilis/genética , Bacillus subtilis/enzimologia , Bacillus subtilis/química , Bacillus subtilis/metabolismo , Biocatálise , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/químicaRESUMO
6'-Sialyllactose (6'-SL), the most abundant sialylated human milk oligosaccharide, has attracted attention for its potential application in supplementary infant formulas. Herein, we report a facile strategy to construct a cascade bioreactor for the enzymatic synthesis of 6'-SL by co-immobilizing an enzymatic module consisting of CMP-sialic acid synthase and α-2,6-sialyltransferase into hierarchically porous MIL-53 (HP-MIL-53). The as-prepared HP-MIL-53 showed high enzyme immobilization capacity, reaching 226 mg g-1. Furthermore, the co-immobilized enzymes exhibited higher initial catalytic efficiency, and thermal, pH and storage stability than the free ones. Finally, the 6'-SL yield remained >80% after 13 cycles of use. We expect that HP-MIL-53 would have potential industrial applications in the enzymatic modular synthesis of 6'-SL and other glycans.
Assuntos
Enzimas Imobilizadas , Sialiltransferases , Enzimas Imobilizadas/química , Enzimas Imobilizadas/metabolismo , Sialiltransferases/metabolismo , Porosidade , Humanos , Oligossacarídeos/química , Oligossacarídeos/metabolismo , Oligossacarídeos/biossíntese , N-Acilneuraminato Citidililtransferase/metabolismo , N-Acilneuraminato Citidililtransferase/química , Reatores Biológicos , Leite Humano/química , Leite Humano/metabolismo , Lactose/química , Lactose/análogos & derivados , Lactose/metabolismo , Concentração de Íons de Hidrogênio , beta-D-Galactosídeo alfa 2-6-SialiltransferaseRESUMO
Carbimazole has disadvantages on different body organs, especially the thyroid gland and, rarely, the adrenal glands. Most studies have not suggested any solution or medication for ameliorating the noxious effects of drugs on the glands. Our study focused on the production of xylooligosaccharide (XOS), which, when coadministered with carbimazole, relieves the toxic effects of the drug on the adrenal glands. In addition to accelerating the regeneration of adrenal gland cells, XOS significantly decreases the oxidative stress caused by obesity. This XOS produced by Aspergillus terreus xylanase was covalently immobilized using microbial Scleroglucan gel beads, which improved the immobilization yield, efficiency, and operational stability. Over a wide pH range (6-7.5), the covalent immobilization of xylanase on scleroglucan increased xylanase activity compared to that of its free form. Additionally, the reaction temperature was increased to 65 °C. However, the immobilized enzyme demonstrated superior thermal stability, sustaining 80.22% of its original activity at 60 °C for 120 min. Additionally, the full activity of the immobilized enzyme was sustained after 12 consecutive cycles, and the activity reached 78.33% after 18 cycles. After 41 days of storage at 4 °C, the immobilized enzyme was still active at approximately 98%. The immobilized enzyme has the capability to produce xylo-oligosaccharides (XOSs). Subsequently, these XOSs can be coadministered alongside carbimazole to mitigate the adverse effects of the drug on the adrenal glands. In addition to accelerating the regeneration of adrenal gland cells, XOS significantly decreases the oxidative stress caused by obesity.