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1.
Adv Sci (Weinh) ; : e2400874, 2024 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-38760899

RESUMO

Various electrocatalysts are extensively examined for their ability to selectively produce desired products by electrochemical CO2 reduction reaction (CO2RR). However, an efficient CO2RR electrocatalyst doesn't ensure an effective co-catalyst on the semiconductor surface for photoelectrochemical CO2RR. Herein, Bi2S3 nanorods are synthesized and electrochemically reduced to Bi nanoplates that adhere to the substrates for application in the electrochemical and photoelectrochemical CO2RR. Compared with commercial-Bi, the Bi2S3-derived Bi (S-Bi) nanoplates on carbon paper exhibit superior electrocatalytic activity and selectivity for formate (HCOO-) in the electrochemical CO2RR, achieving a Faradaic efficiency exceeding 93%, with minimal H2 production over a wide potential range. This highly selective S-Bi catalyst is being employed on the Si photocathode to investigate the behavior of electrocatalysts during photoelectrochemical CO2RR. The strong adhesion of the S-Bi nanoplates to the Si nanowire substrate and their unique catalytic properties afford exceptional activity and selectivity for HCOO- under simulated solar irradiation. The selectivity observed in electrochemical CO2RR using the S-Bi catalyst correlates with that seen in the photoelectrochemical CO2RR system. Combined pulsed potential methods and theoretical analyses reveal stabilization of the OCHO* intermediate on the S-Bi catalyst under specific conditions, which is critical for developing efficient catalysts for CO2-to-HCOO- conversion.

2.
Int J Biol Macromol ; 269(Pt 2): 131834, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38688341

RESUMO

The amylosucrase (ASase, EC 2.4.1.4) utilizes sucrose as the sole substrate to catalyze multifunctional reactions. It can naturally synthesize α-1,4-linked glucans such as amylose as well as sucrose isomers with more favorable properties than sucrose with a lower intestinal digestibility and non-cariogenic properties. The amino acid sequence of the asase gene from Deinococcus cellulosilyticus (DceAS) exhibits low homology with those of other ASases from other Deinococcus species. In this study, we cloned and expressed DceAS and demonstrated its high activity at pH 6 and pH 8 and maintained stability. It showed higher polymerization activity at pH 6 than at pH 8, but similar isomerization activity and produced more turanose and trehalulose at pH 6 than at pH 8 and produced more isomaltulose at pH 8. Furthermore, the molecular weight of DceAS was 226.6 kDa at pH 6 and 145.5 kDa at pH 8, indicating that it existed as a trimer and dimer, respectively under those conditions. Additionally, circular dichroism spectra showed that the DceAS secondary structure was different at pH 6 and pH 8. These differences in reaction products at different pHs can be harnessed to naturally produce sucrose alternatives that are more beneficial to human health.


Assuntos
Deinococcus , Glucosiltransferases , Glucosiltransferases/química , Glucosiltransferases/genética , Glucosiltransferases/metabolismo , Deinococcus/enzimologia , Deinococcus/genética , Concentração de Íons de Hidrogênio , Isomaltose/metabolismo , Isomaltose/química , Isomaltose/análogos & derivados , Sequência de Aminoácidos , Estabilidade Enzimática , Clonagem Molecular , Peso Molecular , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Sacarose/metabolismo , Especificidade por Substrato , Cinética , Estrutura Secundária de Proteína , Dissacarídeos
3.
Food Chem ; 448: 139182, 2024 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-38569413

RESUMO

Amylosucrase (ASase) efficiently biosynthesizes α-glucoside using flavonoids as acceptor molecules and sucrose as a donor molecule. Here, ASase from Deinococcus wulumuqiensis (DwAS) biosynthesized more naringenin α-glucoside (NαG) with sucrose and naringenin as donor and acceptor molecules, respectively, than other ASases from Deinococcus sp. The biotransformation rate of DwAS to NαG was 21.3% compared to 7.1-16.2% for other ASases. Docking simulations showed that the active site of DwAS was more accessible to naringenin than those of others. The 217th valine in DwAS corresponded to the 221st isoleucine in Deinococcus geothermalis AS (DgAS), and the isoleucine possibly prevented naringenin from accessing the active site. The DwAS-V217I mutant had a significantly lower biosynthetic rate of NαG than DwAS. The kcat/Km value of DwAS with naringenin as the donor was significantly higher than that of DgAS and DwAS-V217I. In addition, NαG inhibited human intestinal α-glucosidase more efficiently than naringenin.


Assuntos
Proteínas de Bactérias , Biotransformação , Deinococcus , Flavanonas , Glucosídeos , Glucosiltransferases , Inibidores de Glicosídeo Hidrolases , Flavanonas/metabolismo , Flavanonas/química , Deinococcus/enzimologia , Deinococcus/metabolismo , Deinococcus/química , Deinococcus/genética , Glucosiltransferases/metabolismo , Glucosiltransferases/química , Glucosiltransferases/genética , Inibidores de Glicosídeo Hidrolases/química , Inibidores de Glicosídeo Hidrolases/metabolismo , Inibidores de Glicosídeo Hidrolases/farmacologia , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Glucosídeos/metabolismo , Glucosídeos/química , Simulação de Acoplamento Molecular , Cinética , alfa-Glucosidases/metabolismo , alfa-Glucosidases/química
4.
Biomacromolecules ; 25(3): 2024-2032, 2024 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-38393758

RESUMO

α-Glucan microparticles (GMPs) have significant potential as high-value biomaterials in various industries. This study proposes a bottom-up approach for producing GMPs using four amylosucrases from Bifidobacterium sp. (BASs). The physicochemical characteristics of these GMPs were analyzed, and the results showed that the properties of the GMPs varied depending on the type of enzymes used in their synthesis. As common properties, all GMPs exhibited typical B-type crystal patterns and poor colloidal dispersion stability. Interestingly, differences in the physicochemical properties of GMPs were generated depending on the synthesis rate of linear α-glucan by the enzymes and the degree of polymerization (DP) distribution. Consequently, we found differences in the properties of GMPs depending on the DP distribution of linear glucans prepared with four BASs. Furthermore, we suggest that precise control of the type and characteristics of the enzymes provides the possibility of producing GMPs with tailored physicochemical properties for various industrial applications.


Assuntos
Bifidobacterium , Glucanos , Guanosina Monofosfato , Tionucleotídeos , Glucanos/química , Glucosiltransferases
5.
Biomolecules ; 13(8)2023 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-37627337

RESUMO

The diamondback moth is a detrimental insect pest of brassicaceous crops which was among the first crop insects to be reported as DDT resistant. It has since proven to be significantly resistant to nearly every synthetic insecticide used in the field in many crucifer-producing regions. Due to insecticide control failures in some parts of the world, economically viable crucifer production is now all but impossible. As a result, there has been an increasing effort to identify new compounds with strong pesticidal activity. Cantharidin is one such compound that has been shown to be highly effective against a variety of insect pests. However, its chemical synthesis and potential toxicity to non-target organisms have been a major source of concern. Herein, using rational design approaches, a new series of cantharidin-based verbenone derivatives were synthesized and evaluated for their insecticidal activities against the diamondback moth. Among different compounds screened, compounds 6a, 6h, 6i, and 6q emerged as the most potent compounds exhibiting 100% mortality at a concentration of 100 mg/L after four days. These compounds demonstrated a good anti-feeding effect against the diamondback moth on cabbage leaves. Subsequently, a 3D QSAR study was carried out to identify the key structural features of the synthesized compounds and their correlation with insecticidal activity.


Assuntos
Inseticidas , Inseticidas/farmacologia , Cantaridina/farmacologia , Relação Quantitativa Estrutura-Atividade , Monoterpenos Bicíclicos
6.
Food Sci Biotechnol ; 32(9): 1299, 2023 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-37362806

RESUMO

[This corrects the article DOI: 10.1007/s10068-019-00686-6.].

7.
Food Sci Biotechnol ; 32(6): 749-768, 2023 May.
Artigo em Inglês | MEDLINE | ID: mdl-37041815

RESUMO

Microbial biocatalysts are evolving technological tools for glycosylation research in food, feed and pharmaceuticals. Advances in bioengineered Leloir and non-Leloir carbohydrate-active enzymes allow for whole-cell biocatalysts to curtail production costs of purified enzymes while enhancing glucan synthesis through continued enzyme expression. Unlike sugar nucleotide-dependent Leloir glycosyltransferases, non-Leloir enzymes require inexpensive sugar donors and can be designed to match the high value, yield and selectivity of the former. This review addresses the current state of bacterial cell-based production of glucans and glycoconjugates via transglycosylation, and describes how alterations made to microbial hosts to surpass purified enzymes as the preferred mode of catalysis are steadily being acquired through genetic engineering, rational design and process optimization. A comprehensive exploration of relevant literature has been summarized to describe whole-cell biocatalysis in non-Leloir glycosylation reactions with various donors and acceptors, and the characterization, application and latest developments in the optimization of their use.

8.
Food Sci Biotechnol ; 32(4): 565-575, 2023 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-36911326

RESUMO

Amylosucrase can increase the amount of resistant starch (RS) in starch by transferring glucose from sucrose to amylopectin. Here, rice starch was modified using amylosucrase from Deinococcus geothermalis (DgAS). DgAS-modified rice starch (DMRS) increased the side-chain length of amylopectin and appeared in the form of B-type crystals. In vitro digestion analyses revealed that DMRS had a higher RS contents and lower digestion rate than native rice starch. When high-fat diet (HFD)-induced C57BL/6 mice were orally administered DMRS, body weight and white fat tissues of DMRS-fed HFD mice were not significantly different. However, serum leptin and glucose levels were significantly decreased and serum glucagon like peptide-1was increased in these mice. The cecal microbiome in DMRS-fed HFD mice was identified to investigate the role of DMRS in gut microbiota regulation. DMRS supplementation increased the relative abundance of Bacteroides, Faecalibaculum, and Ruminococcus in mouse gut microbiota. Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-022-01238-1.

9.
Front Microbiol ; 13: 1016675, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36274706

RESUMO

The 4-α-glucanotransferase (4-α-GTase or amylomaltase) is an essential enzyme in maltodextrin metabolism. Generally, most bacterial 4-α-GTase is classified into glycoside hydrolase (GH) family 77. However, hyperthermophiles have unique 4-α-GTases belonging to GH family 57. These enzymes are the main amylolytic protein in hyperthermophiles, but their mode of action in maltooligosaccharide utilization is poorly understood. In the present study, we investigated the catalytic properties of 4-α-GTase from the hyperthermophile Pyrococcus sp. ST04 (PSGT) in the presence of maltooligosaccharides of various lengths. Unlike 4-α-GTases in GH family 77, GH family 57 PSGT produced maltotriose in the early stage of reaction and preferred maltose and maltotriose over glucose as the acceptor. The kinetic analysis showed that maltotriose had the lowest KM value, which increased amylose degradation activity by 18.3-fold. Structural models of PSGT based on molecular dynamic simulation revealed two aromatic amino acids interacting with the substrate at the +2 and +3 binding sites, and the mutational study demonstrated they play a critical role in maltotriose binding. These results clarify the mode of action in carbohydrate utilization and explain acceptor binding mechanism of GH57 family 4-α-GTases in hyperthermophilic archaea.

10.
Food Sci Biotechnol ; 31(9): 1179-1188, 2022 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-35919361

RESUMO

Amylosucrase from Neisseria polysaccharea (NpAS) synthesizes α-1,4 glucan polymer from sucrose. In this study, we coated various botanical sources of raw starch with an α-glucan layer generated by NpAS to improve physicochemical properties. Field-emission scanning electron microscopy demonstrated that all surfaces of the starch granules were successfully coated via the NpAS reaction. X-ray diffraction analysis revealed that the crystallinity decreased and the crystal pattern changed to C-type as an amylose layer formed around the surface of the starch granules. Based on rapid viscosity and differential scanning calorimetry analyses, the gelatinization resistance of the α-glucan-coated starch increased owing to decreased viscosity and increased melting temperature. Therefore, the α-glucans coated the starches by enzymatic reactions of various botanical sources; these have applicability in the food and starch industries owing to various physicochemical properties such as enhanced thermostability. Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-022-01113-z.

11.
ACS Appl Mater Interfaces ; 14(32): 36557-36569, 2022 Aug 17.
Artigo em Inglês | MEDLINE | ID: mdl-35917313

RESUMO

Enhancement of redox-reversibility in electroactive species has been studied because of fundamental interest and their importance for energy storage systems. Various electroactive molecules suffer from redox-irreversible behavior, and this is a critical reason for their exclusion as redox electrolytes in energy storage systems. In this article, we fully demonstrated that ascorbic acid (ASC), which is an abundant but redox-irreversible molecule, can become redox-reversible when it is confined in microporous carbon regimes. From a theoretical perspective, redox-reversibility in an electrochemical reaction coupled with an irreversible chemical process can be greatly enhanced due to kinetic acceleration toward the inverse direction of the chemical reaction by accumulation of products in the nanoconfined regime. However, the kinetic acceleration in a nanoconfined domain shows limitations for enhancing the redox-reversibility, which indicates that stabilization of the species undergoing an irreversible chemical process is another important factor for redox-reversibility enhancement. The origin of nanoporous confinement of ASC and its enhanced redox-reversibility was rationalized by molecular dynamics simulations. We found that ASC-clusters of a fully protonated ASC and its conjugated base formed inside carbon pores, which would be a main driving force for its confinement in microporous carbon networks. Lastly, we demonstrated a prototype energy storage device using redox-reversible ASC in microporous carbon as the half electrode, which shows the feasibility of ASC as a possible redox electrolyte in an aqueous energy storage system.

12.
Adv Sci (Weinh) ; 9(20): e2201491, 2022 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-35501291

RESUMO

Electrochemical carbon dioxide reduction is a mild and eco-friendly approach for CO2 mitigation and producing value-added products. For selective electrochemical CO2 reduction, single-crystalline Au particles (octahedron, truncated-octahedron, and sphere) are synthesized by consecutive growth and chemical etching using a polydiallyldimethylammonium chloride (polyDDA) surfactant, and are surface-functionalized. Monodisperse, single-crystalline Au nanoparticles provide an ideal platform for evaluating the Au surface as a CO2 reduction catalyst. The polyDDA-Au cathode affords high catalytic activity for CO production, with >90% Faradaic efficiency over a wide potential range between -0.4 and -1.0 V versus RHE, along with high durability owing to the consecutive interaction between dimethylammonium and chloride on the Au surface. The influence of polyDDA on the Au particles, and the origins of the enhanced selectivity and stability are fully investigated using theoretical studies. Chemically adsorbed polyDDA is consecutively affected the initial adsorption of CO2 and the stability of the *CO2 , *COOH, and *CO intermediates during continuous CO2 reduction reaction. The polyDDA functionalization is extended to improving the CO Faradaic efficiency of other metal catalysts such as Ag and Zn, indicating its broad applicability for CO2 reduction.

13.
Food Sci Biotechnol ; 31(2): 231-241, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-35186353

RESUMO

Resistant starch (RS) in the diet reaches the large intestine and is fermented by the gut microbiota, providing beneficial effects on human health. The human gut bacterium FMB-CY1 was isolated and identified as a new species closest to Ruminococcus bromii. Ruminococcus sp. FMB-CY1 completely degraded RS including commercial RS types 2, 3, and 4, and generated glucose and maltose; however, it did not assimilate glucose. Genome analysis revealed 15 amylolytic enzymes (Amy) present in FMB-CY1. The evolutionary trees revealed that the Amys were well divided each other. All Amys (4, 9, 10, 12, and 16) containing cohesin and/or dockerin and scaffolding proteins known to be involved in constituting the amylosome, were identified. A new species of Ruminococcus, strain FMB-CY1, was considered to have the ability to form amylosomes for the degradation of RS. This new RS-degrading Ruminococcus species provides insights into the mechanism(s) underlying RS degradation in the human gut. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10068-021-01027-2.

14.
Int J Biol Macromol ; 193(Pt B): 1340-1349, 2021 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-34740684

RESUMO

A putative type II pullulanase gene, pulP, was identified in Bifidobacterium adolescentis P2P3. PulP possesses an α-amylase domain at the N-terminus and a pullulanase type I domain at the C-terminus, as well as three carbohydrate-binding modules (one CBM25 and two CBM41s) between them. The native PulP and four truncated mutant recombinant proteins (PulPΔCΔP, PulPΔP, PulPΔAΔC, and PulPΔA), in which each of the two catalytic domains and/or the CBMs were deleted, were produced in Escherichia coli and their specific properties were characterized. The removal of either catalytic domain abolished the corresponding catalytic activity of the wild-type enzyme. Deletion of the C-terminal domain resulted in a drastic decrease in the optimal temperature and thermostability, indicating that the pullulanase domain might be related to the temperature dependency of the enzyme. In addition, the elimination of the CBMs in the mutant proteins led to a loss of binding affinity toward raw substrates as well as the loss of their hydrolysis activities compared to the wild-type enzyme. HPAEC and TLC analyses proved that PulP and its mutants could hydrolyze α-glucans into maltotriose as their main product. These results suggest that PulP may play an important role in α-glucan metabolism in B. adolescentis P2P3.


Assuntos
Proteínas de Bactérias/metabolismo , Bifidobacterium adolescentis/metabolismo , Microbioma Gastrointestinal/fisiologia , Glicosídeo Hidrolases/metabolismo , Amido Resistente/metabolismo , Amido/metabolismo , Metabolismo dos Carboidratos/fisiologia , Catálise , Escherichia coli/metabolismo , Glucanos/metabolismo , Hidrólise , Proteínas Recombinantes/metabolismo , alfa-Amilases/metabolismo
15.
Food Sci Biotechnol ; 30(2): 267-276, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33732517

RESUMO

Amylosucrase (ASase, E.C. 2.4.1.4) is a powerful transglycosylation enzyme that can transfer glucose from sucrose to the hydroxyl (-OH) group of various compounds. In this study, recombinant ASases from Deinococcus geothermalis (DgAS) and Bifidobacterium thermophilum (BtAS) were used to synthesize biosurfactants based on the computational analysis of predicted docking simulations. Successful predictions of the binding affinities, conformations, and three-dimensional structures of three surfactants were computed from receptor-ligand binding modes. DgAS and BtAS were effective in the synthesis of biosurfactants from glyceryl caprylate, glyceryl caprate, and polyglyceryl-2 caprate. The results of the transglycosylation reaction were consistent for both ASases, with glyceryl caprylate acceptor showing the highest concentration, as confirmed by thin layer chromatography. Furthermore, the transglycosylation reactions of DgAS were more effective than those of BtAS. Among the three substrates, glyceryl caprylate glycoside and glyceryl caprate glycoside were successfully purified by liquid chromatography-mass spectrometry (LC-MS) with the corresponding molecular weights.

16.
Genomics ; 113(1 Pt 2): 647-653, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33010389

RESUMO

1-Deoxynojirumycin (1-DNJ) is a representative iminosugar with α-glucosidase inhibition (AGI) activity. In this study, the full genome sequencing of 1-DNJ-producing Bacillus velezensis K26 was performed. The genome consists of a circular chromosome (4,047,350 bps) with two types of putative virulence factors, five antibiotic resistance genes, and seven secondary metabolite biosynthetic gene clusters. Genomic analysis of a wide range of Bacillus species revealed that a 1-DNJ biosynthetic gene cluster was commonly present in four Bacillus species (B. velezensis, B. pseudomycoides, B. amyloliquefaciens, and B. atrophaeus). In vitro experiments revealed that the increased mRNA expression levels of the three 1-DNJ biosynthetic genes were closely related to increased AGI activity. Genomic comparison and alignment of multiple gene sequences indicated the conservation of the 1-DNJ biosynthetic gene cluster in each Bacillus species. This genomic analysis of Bacillus species having a 1-DNJ biosynthetic gene cluster could provide a basis for further research on 1-DNJ-producing bacteria.


Assuntos
Bacillus/genética , Genes Bacterianos , Glucosamina/análogos & derivados , 1-Desoxinojirimicina , Bacillus/classificação , Bacillus/metabolismo , Glucosamina/biossíntese , Glucosamina/genética , Família Multigênica , Filogenia , Homologia de Sequência
17.
J Microbiol Biotechnol ; 31(1): 63-69, 2021 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-33148942

RESUMO

Carotenoids, which have biologically beneficial effects and occur naturally in microorganisms and plants, are pigments widely applied in the food, cosmetics and pharmaceutical industries. The compound 4,4'-diaponeurosporene is a C30 carotenoid produced by some Lactobacillus species, and Lactobacillus plantarum is the main species producing it. In this study, the antioxidant activity of 4,4'-diaponeurosporene extracted from L. plantarum subsp. plantarum KCCP11226 was examined. Maximum carotenoid content (0.74 ± 0.2 at A470) was obtained at a relatively low temperature (20°C). The DPPH radical scavenging ability of 4,4'-diaponeurosporene (1 mM) was approximately 1.7-fold higher than that of butylated hydroxytoluene (BHT), a well-known antioxidant food additive. In addition, the ABTS radical scavenging ability was shown to be 2.3- to 7.5-fold higher than that of BHT at the range of concentration from 0.25 mM to 1 mM. The FRAP analysis confirmed that 4,4'- diaponeurosporene (0.25 mM) was able to reduce Fe3+ by 8.0-fold higher than that of BHT. Meanwhile, 4,4'-diaponeurosporene has been confirmed to be highly resistant to various external stresses (acid/bile, high temperature, and lysozyme conditions). In conclusion, L. plantarum subsp. plantarum KCCP11226, which produces 4,4'-diaponeurosporene as a functional antioxidant, may be a potentially useful strain for the development of functional probiotic industries.


Assuntos
Antioxidantes/química , Carotenoides/metabolismo , Lactobacillus/metabolismo , Triterpenos/metabolismo , Farmacorresistência Bacteriana , Muramidase , Probióticos , Temperatura
18.
Sensors (Basel) ; 20(24)2020 Dec 12.
Artigo em Inglês | MEDLINE | ID: mdl-33322805

RESUMO

This paper proposes a new test method of detecting the presence of impulsive noise based on a complementary cumulative density function (CCDF). Impulsive noise severely degrades performance of communication systems and the conventional Kolmogorov-Smirnov (K-S) test may not perform well, because the test does not consider the characteristics of impulsive noise. In order to detect the presence of impulsive noise reliably, the CCDF of measurement samples is analyzed and compared with the CCDF of additive white Gaussian noise to find the difference between those CCDFs. Due to the nature of heavy-tails in impulsive noise, only the maximum difference may not be sufficient for the accurate detection of impulsive noise. Therefore, the proposed method applies the test hypothesis using the weighted sum of all the differences between those CCDFs. Simulation results justify that the proposed test is more robust and provides lower miss detection probability than the K-S test in the presence of impulsive noise.

19.
Int J Biol Macromol ; 161: 389-397, 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-32479932

RESUMO

Resistant starch (RS) is a complex prebiotic carbohydrate beneficial to the human gut. In the present study, four genes encoding for putative amylolytic enzymes, likely to be responsible for RS-degradation, were identified in the genome of Bifidobacterium adolescentis P2P3 by comparative genomic analysis. Our results showed that only three enzymes (RSD1, RSD2, and RSD3) exhibited non-gelatinized high amylose corn starch (HACS)-degrading activity in addition to typical α-amylase activity. These three RS-degrading enzymes (RSD) were composed of multiple domains, including signal peptide, catalytic domain, carbohydrate binding domains, and putative cell wall-anchoring domains. Typical catalytic domains were conserved by exhibiting seven typical conserved regions (I-VII) found mostly in α-amylases. Analysis of enzymatic activity revealed that RSD2 displayed stronger activity toward HACS-granules than RSD1 and RSD3. Comparative genomics in combination with enzymatic experiments confirmed that RSDs might be the key enzymes used by RS-degrading bifidobacteria to degrade RS in a particular ecological niche, such as the human gut.


Assuntos
Amilases/metabolismo , Bifidobacterium adolescentis/enzimologia , Microbioma Gastrointestinal , Amido Resistente/metabolismo , Sequência de Aminoácidos , Amilases/química , Bifidobacterium/classificação , Bifidobacterium/enzimologia , Bifidobacterium/genética , Bifidobacterium adolescentis/classificação , Bifidobacterium adolescentis/genética , Biologia Computacional/métodos , Genoma Bacteriano , Humanos , Hidrólise , Filogenia
20.
J Microbiol Biotechnol ; 30(9): 1436-1442, 2020 Sep 28.
Artigo em Inglês | MEDLINE | ID: mdl-32522959

RESUMO

Amylosucrase (ASase, E.C. 2.4.1.4) is capable of efficient glucose transfer from sucrose, acting as the sole donor molecule, to various functional acceptor compounds, such as polyphenols and flavonoids. An ASase variant from Deinococcus geothermalis, in which the 226th alanine is replaced with asparagine (DgAS-A226N), shows increased polymerization activity due to changes in the flexibility of the loop near the active site. In this study, we further investigated how the mutation modulates the enzymatic activity of DgAS using molecular dynamics and docking simulations to evaluate interactions between the enzyme and phenolic compounds. The computational analysis revealed that the A226N mutation could induce and stabilize structural changes near the substratebinding site to increase glucose transfer efficiency to phenolic compounds. Kinetic parameters of DgAS-A226N and WT DgAS were determined with sucrose and 4-methylumbelliferone (MU) as donor and acceptor molecules, respectively. The kcat/Km value of DgAS-A226N with MU (6.352 mM-1min-1) was significantly higher than that of DgAS (5.296 mM-1min-1). The enzymatic activity was tested with a small phenolic compound, hydroquinone, and there was a 1.4-fold increase in α-arbutin production. From the results of the study, it was concluded that DgAS-A226N has improved acceptor specificity toward small phenolic compounds by way of stabilizing the active conformation of these compounds.


Assuntos
Deinococcus/enzimologia , Glucosiltransferases/química , Glucosiltransferases/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Deinococcus/genética , Glucose/metabolismo , Glucosiltransferases/genética , Hidroquinonas , Cinética , Simulação de Acoplamento Molecular , Sacarose/metabolismo
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