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1.
Int J Mol Sci ; 22(19)2021 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-34638570

RESUMO

The microbial biodegradation of new PLA and PCL materials containing birch tar (1-10% v/v) was investigated. Product of dry distillation of birch bark (Betula pendula Roth) was added to polymeric materials to obtain films with antimicrobial properties. The subject of the study was the course of enzymatic degradation of a biodegradable polymer with antibacterial properties. The results show that the type of the material, tar concentration, and the environment influenced the hydrolytic activity of potential biofilm degraders. In the presence of PCL films, the enzyme activities were higher (except for α-D-glucosidase) compared to PLA films. The highest concentration of birch tar (10% v/v) decreased the activity of hydrolases produced by microorganisms to the most significant extent; however, SEM analysis showed the presence of a biofilm even on plastics with the highest tar content. Based on the results of the biological oxygen demand (BOD), the new materials can be classified as biodegradable but, the biodegradation process was less efficient when compared to plastics without the addition of birch tar.


Assuntos
Anti-Infecciosos/química , Betula/química , Plásticos Biodegradáveis/química , Poliésteres/química , Alcatrões/química , Aminopeptidases/metabolismo , Anti-Infecciosos/farmacologia , Bactérias/efeitos dos fármacos , Bactérias/enzimologia , Betula/microbiologia , Plásticos Biodegradáveis/farmacologia , Biofilmes , Análise da Demanda Biológica de Oxigênio , Destilação , Ensaios Enzimáticos , Esterases/metabolismo , Lipase/metabolismo , Casca de Planta/química , Casca de Planta/microbiologia , Poliésteres/metabolismo , Alcatrões/farmacologia , alfa-Glucosidases/metabolismo , beta-Glucosidase/metabolismo
2.
Molecules ; 26(15)2021 Jul 27.
Artigo em Inglês | MEDLINE | ID: mdl-34361670

RESUMO

When wine grapes are exposed to smoke, there is a risk that the resulting wines may possess smoky, ashy, or burnt aromas, a wine flaw known as smoke taint. Smoke taint occurs when the volatile phenols (VPs) largely responsible for the aroma of smoke are transformed in grape into a range of glycosides that are imperceptible by smell. The majority of VP-glycosides described to date are disaccharides possessing a reducing ß-d-glucopyranosyl moiety. Here, a two-part experiment was performed to (1) assess the stability of 11 synthesized VP-glycosides towards general acid-catalyzed hydrolysis during aging, and (2) to examine whether yeast strains differed in their capacity to produce free VPs both from these model glycosides as well as from grapes that had been deliberately exposed to smoke. When fortified into both model and real wine matrices at 200 ng/g, all VP-disaccharides were stable over 12 weeks, while (42-50 ng/g) increases in free 4-ethylphenol and p-cresol were detected when these were added to wine as their monoglucosides. Guaiacol and phenol were the most abundantly produced VPs during fermentation, whether originating from natural VP-precursors in smoked-exposed Pinot Noir must, or due to fortification with synthetic VP-glycosides. Significant yeast strain-specific differences in glycolytic activities were observed for phenyl-ß-d-glycopyranoside, with two strains (RC212 and BM45) being unable to hydrolyze this model VP, albeit both were active on the guaiacyl analogue. Thus, differences in Saccharomyces cerevisiae ß-glucosidase activity appear to be influenced by the VP moiety.


Assuntos
Fermentação , Frutas/metabolismo , Glicosídeos/metabolismo , Odorantes/análise , Fenol/metabolismo , Saccharomyces cerevisiae/enzimologia , Fumaça/efeitos adversos , Vitis/metabolismo , Compostos Orgânicos Voláteis/metabolismo , Vinho/análise , Cresóis/metabolismo , Guaiacol/metabolismo , Fenóis/metabolismo , beta-Glucosidase/metabolismo
3.
Food Microbiol ; 100: 103859, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34416959

RESUMO

Beta-glucosidase is an important enzyme for the hydrolysis of grape glycosides in the course of winemaking. Yeasts are the main producers of ß-glucosidase in winemaking, therefore play an important role in determining wine aroma and flavour. This article discusses common methods for ß-glucosidase evaluation, the ß-glucosidase activity of different Saccharomyces and non- Saccharomyces yeasts and the influences of winemaking conditions, such as glucose and ethanol concentration, low pH environment, fermentation temperature and SO2 level, on their activity. This review further highlights the roles of ß-glucosidase in promoting the release of free volatile compounds especially terpenes and the modification of wine phenolic composition during the winemaking process. Furthermore, this review proposes future research direction in this area and guides wine professionals in yeast selection to improve wine quality.


Assuntos
Proteínas Fúngicas/metabolismo , Compostos Orgânicos Voláteis/metabolismo , Vinho/análise , Leveduras/enzimologia , beta-Glucosidase/metabolismo , Proteínas Fúngicas/genética , Odorantes/análise , Fenóis/química , Fenóis/metabolismo , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Vitis/química , Vitis/microbiologia , Compostos Orgânicos Voláteis/química , Vinho/microbiologia , Leveduras/genética , Leveduras/metabolismo , beta-Glucosidase/genética
4.
Biochem Biophys Res Commun ; 569: 61-65, 2021 09 10.
Artigo em Inglês | MEDLINE | ID: mdl-34229124

RESUMO

For the beneficial pharmacological properties of isoflavonoids and their related glycoconjugates, there is increasingly interest in their enzymatic conversion. In this study, a novel ß-glucosidase gene isolated from metagenomic library of mangrove sediment was cloned and overexpressed in Escherichia coli BL21(DE3). The purified recombination ß-glucosidase, designated as r-Bgl66, showed high catalytic activity for soy isoflavone glycosides. It converted soy isoflavone flour extract with the productivities of 0.87 mM/h for daidzein, 0.59 mM/h for genistein and 0.42 mM/h for glycitein. The kcat/Km values for daidzin, genistin and glycitin were 208.73, 222.37 and 288.07 mM-1 s-1, respectively. In addition, r-Bgl66 also exhibited the characteristic of glucose-tolerance, and the inhibition constant Ki was 471.4 mM. These properties make it a good candidate in the enzymatic hydrolysis of soy isoflavone glycosides. This study also highlights the utility of metagenomic approach in discovering novel ß-glucosidase for soy isoflavone glycosides hydrolysis.


Assuntos
Avicennia/crescimento & desenvolvimento , Glicosídeos/metabolismo , Isoflavonas/metabolismo , Metagenoma/genética , Microbiologia do Solo , beta-Glucosidase/metabolismo , Biocatálise/efeitos dos fármacos , Eletroforese em Gel de Poliacrilamida , Biblioteca Gênica , Sedimentos Geológicos/microbiologia , Glucose/metabolismo , Glucose/farmacologia , Hidrólise , Cinética , Proteínas Recombinantes/metabolismo , Soja/metabolismo , beta-Glucosidase/genética , beta-Glucosidase/isolamento & purificação
5.
Int J Mol Sci ; 22(14)2021 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-34299210

RESUMO

Conjugation of phytohormones with glucose is a means of modulating their activities, which can be rapidly reversed by the action of ß-glucosidases. Evaluation of previously characterized recombinant rice ß-glucosidases found that nearly all could hydrolyze abscisic acid glucose ester (ABA-GE). Os4BGlu12 and Os4BGlu13, which are known to act on other phytohormones, had the highest activity. We expressed Os4BGlu12, Os4BGlu13 and other members of a highly similar rice chromosome 4 gene cluster (Os4BGlu9, Os4BGlu10 and Os4BGlu11) in transgenic Arabidopsis. Extracts of transgenic lines expressing each of the five genes had higher ß-glucosidase activities on ABA-GE and gibberellin A4 glucose ester (GA4-GE). The ß-glucosidase expression lines exhibited longer root and shoot lengths than control plants in response to salt and drought stress. Fusions of each of these proteins with green fluorescent protein localized near the plasma membrane and in the apoplast in tobacco leaf epithelial cells. The action of these extracellular ß-glucosidases on multiple phytohormones suggests they may modulate the interactions between these phytohormones.


Assuntos
Ácido Abscísico/farmacologia , Ésteres/química , Glucose/metabolismo , Oryza/enzimologia , Proteínas de Plantas/metabolismo , beta-Glucosidase/metabolismo , Ácido Abscísico/química , Arabidopsis/efeitos dos fármacos , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Secas , Giberelinas/farmacologia , Hidrólise , Família Multigênica , Reguladores de Crescimento de Plantas/farmacologia , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/efeitos dos fármacos , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Plantas Geneticamente Modificadas/metabolismo , Tabaco/efeitos dos fármacos , Tabaco/crescimento & desenvolvimento , Tabaco/metabolismo , beta-Glucosidase/genética
6.
Int J Mol Sci ; 22(13)2021 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-34202076

RESUMO

Mutations in GBA1, the gene encoding glucocerebrosidase, are common genetic risk factors for Parkinson disease (PD). While the mechanism underlying this relationship is unclear, patients with GBA1-associated PD often have an earlier onset and faster progression than idiopathic PD. Previously, we modeled GBA1-associated PD by crossing gba haploinsufficient mice with mice overexpressing a human mutant α-synuclein transgene (SNCAA53T), observing an earlier demise, shorter life span and faster symptom progression, although behavioral testing was not performed. To assess whether gba+/-//SNCAA53T mice exhibit a prodromal behavioral phenotype, we studied three cardinal PD features: olfactory discrimination, memory dysfunction, and motor function. The longitudinal performance of gba+/-//SNCAA53T (n = 8), SNCAA53T (n = 9), gba+/- (n = 10) and wildtype (n = 6) mice was evaluated between ages 8 and 23 months using the buried pellet test, novel object recognition test and the beam walk. Fifteen-month-old gba+/-//SNCAA53T mice showed more olfactory and motor deficits than wildtype mice. However, differences between gba+/-//SNCAA53T and SNCAA53T mice generally did not reach statistical significance, possibly due to small sample sizes. Furthermore, while gba haploinsufficiency leads to a more rapid demise, this might not result in an earlier prodromal stage, and other factors, including aging, oxidative stress and epigenetics, may contribute to the more fulminant disease course.


Assuntos
Mutação , Doença de Parkinson/etiologia , Fenótipo , beta-Glucosidase/genética , Alelos , Animais , Modelos Animais de Doenças , Feminino , Genótipo , Humanos , Masculino , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Doença de Parkinson/metabolismo , Doença de Parkinson/patologia , beta-Glucosidase/metabolismo
7.
J Chromatogr A ; 1653: 462388, 2021 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-34280789

RESUMO

In this study, an enzyme immobilization method for the effective biotransformation of ginsenoside Rb1 to impart activity and stability was developed. Using a hydrolase enzyme model, ß-glucosidase from Aspergillus niger, immobilization within chemically affinity-linked amino-based silica provided an immobilization efficiency 5.86-fold higher than that of free enzyme. Compared with the free enzyme, the immobilized enzyme functioned optimally at a wider pH range and had higher thermostability. The optimum pH for the free and immobilized enzymes was 5.5. The optimal reaction temperature of the immobilized enzyme was 45 °C, which was 5 °C higher than that of the free enzyme. The Michaelis constant (Km) values before and after immobilization were 0.482 mmol•L-1 and 0.387 mmol•L-1, respectively. The catalytic rate (Kcat) for the immobilized and free enzymes was 22.269 mmol•L-1and 8.800 mmol•L-1, respectively, and the catalytic efficiency (Kcat/Km) activity of the immobilized enzyme was 3.30-fold higher than that of the free enzyme. The immobilized enzyme could preserve 97 % of the activity after 45 cycles of repeated use. The high catalytic activity and significant operational stability are beneficial for industrial applications.


Assuntos
Aspergillus niger , Ginsenosídeos , Dióxido de Silício , beta-Glucosidase , Aspergillus niger/metabolismo , Biotransformação , Estabilidade Enzimática , Enzimas Imobilizadas/metabolismo , Ginsenosídeos/química , Ginsenosídeos/metabolismo , Concentração de Íons de Hidrogênio , Dióxido de Silício/química , Temperatura , beta-Glucosidase/metabolismo
8.
J Microbiol Biotechnol ; 31(7): 1035-1043, 2021 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-34226403

RESUMO

Although engineered Saccharomyces cerevisiae fermenting cellobiose is useful for the production of biofuels from cellulosic biomass, cellodextrin accumulation is one of the main problems reducing ethanol yield and productivity in cellobiose fermentation with S. cerevisiae expressing cellodextrin transporter (CDT) and intracellular ß-glucosidase (GH1-1). In this study, we investigated the reason for the cellodextrin accumulation and how to alleviate its formation during cellobiose fermentation using engineered S. cerevisiae fermenting cellobiose. From the series of cellobiose fermentation using S. cerevisiae expressing only GH1-1 under several culture conditions, it was discovered that small amounts of GH1-1 were secreted and cellodextrin was generated through trans-glycosylation activity of the secreted GH1-1. As GH1-1 does not have a secretion signal peptide, non-conventional protein secretion might facilitate the secretion of GH1-1. In cellobiose fermentations with S. cerevisiae expressing only GH1-1, knockout of TLG2 gene involved in non-conventional protein secretion pathway significantly delayed cellodextrin formation by reducing the secretion of GH1-1 by more than 50%. However, in cellobiose fermentations with S. cerevisiae expressing both GH1-1 and CDT-1, TLG2 knockout did not show a significant effect on cellodextrin formation, although secretion of GH1-1 was reduced by more than 40%. These results suggest that the development of new intracellular ß-glucosidase, not influenced by non-conventional protein secretion, is required for better cellobiose fermentation performances of engineered S. cerevisiae fermenting cellobiose.


Assuntos
Celobiose/metabolismo , Celulose/análogos & derivados , Dextrinas/metabolismo , Saccharomyces cerevisiae/metabolismo , beta-Glucosidase/metabolismo , Biocombustíveis , Celulose/metabolismo , Etanol/metabolismo , Fermentação , Glicosilação , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Engenharia Metabólica , Proteínas Qa-SNARE/genética , Proteínas Qa-SNARE/metabolismo , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Via Secretória/genética , beta-Glucosidase/genética
9.
Enzyme Microb Technol ; 148: 109799, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34116753

RESUMO

Most of the presently known ß-glucosidases are sensitive to end-product inhibition by glucose, restricting their potential use in many industrial applications. Identification of novel glucose tolerant ß-glucosidase can prove a pivotal solution to eliminate end-product inhibition and enhance the overall lignocellulosic saccharification process. In this study, a novel gene encoding ß-glucosidase BglNB11 of 1405bp was identified in the genome of Saccharomonospora sp. NB11 and was successfully cloned and heterologously expressed in E. coli BL21 (DE3).The presence of conserved amino acids; NEPW and TENG indicated that BglNB11 belonged to GH1 ß-glucosidases. The recombinant enzyme was purified using a Ni-NTA column, with the molecular mass of 51 kDa, using SDS-PAGE analysis. BglNB11 showed optimum activity at 40 °C and pH 7 and did not require any tested co-factors for activation. The kinetic values, Km, Vmax, kcat, and kcat/Km of purified enzyme were 0.4037 mM, 5735.8 µmol/min/mg, 5042.16 s-1 and 12487.71 s-1 mM-1, respectively. The enzyme was not inhibited by glucose to a concentration of 4 M but was slightly stimulated in the presence of glucose. Molecular docking of BglNB11 with glucose suggested that the relative binding position of glucose in the active site channel might be responsible for modulating end product tolerance and stimulation. ß-glucosidase from BglNB11 is an excellent enzyme with high catalytic efficiency and enhanced glucose tolerance compared to many known glucose tolerant ß-glucosidases. These unique properties of BglNB11 make it a prime candidate to be utilized in many biotechnological applications.


Assuntos
Glucose , beta-Glucosidase , Clonagem Molecular , Estabilidade Enzimática , Escherichia coli/genética , Escherichia coli/metabolismo , Concentração de Íons de Hidrogênio , Simulação de Acoplamento Molecular , Especificidade por Substrato , Temperatura , beta-Glucosidase/genética , beta-Glucosidase/metabolismo
10.
Enzyme Microb Technol ; 148: 109814, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34116760

RESUMO

The ß-glucosidase derived from microorganisms has attracted worldwide interest for their industrial applications, but studies on ß-glucosidases from Oenococcus oeni are rare. In this paper, catalytic mechanism of a novel ß-glucosidase BGL0224 of Oenococcus oeni SD-2a was explored for the first time by kinetic parameters determination, fluorescence spectroscopy and quenching mechanism analysis, molecular dynamics simulation. The results indicated that BGL0224 had universal catalytic effect on different types of glycoside substrates, but the catalytic efficiencies were different. Fluorescence quenching analysis results suggested that the quenching processes between BGL0224 and seven kinds of substrates were predominated by the static quenching mechanism. A reasonable three-dimensional model of BGL0224 was obtained using the crystal structure of E.coli BglA as a template. The analysis results of molecular simulation (RMSD, Rg, RMSF and hydrogen bonding) showed that the composite system 'BGL0224-pNPG' was very stable after 40 ns. The catalytic process of BGL0224 acting on 'p-Nitrophenyl ß-d-glucopyranoside' conformed to the double displacement mechanism. Two glutamic acid residues 'Glu178 and Glu377' played a vital role in the whole catalytic process. Overall, this study gave specific insights on the catalytic mechanism of BGL0224, which was of great significance for developing its potential applications in food industry.


Assuntos
Oenococcus , beta-Glucosidase , Catálise , Cinética , Simulação de Acoplamento Molecular , Oenococcus/metabolismo , beta-Glucosidase/metabolismo
11.
J Ind Microbiol Biotechnol ; 48(5-6)2021 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-34124750

RESUMO

Alkyl glycosides are well-characterized nonionic surfactants, and can be prepared by transglycosylation reactions with retaining GH1 glycosidases being normally used for this purpose. The produced alkyl glycosides can also be hydrolyzed by the glycosidase, and hence, the yields of alkyl glycosides can be too low for industrial use. To improve the transglycosylation-to-hydrolysis ratio for a ß-glucosidase from Thermotoga maritima (TmBglA) for the synthesis of alkyl glycoside, six mutants (N222F, N223C, N223Q, G224A, Y295F, and F414S) were produced. N222F, N223C, N223Q, G224A improved catalytic activity, F295Y and F414S are hydrolytically crippled with p-nitrophenol-ß-d-glucopyranoside (pNPG) as substrate with an 85 and 70-fold decrease in apparent kcat, respectively; N222F shows the highest kcat/km value for pNPG. The substrate selectivity altered from pNPG to pNP-ß-d-fucoside for N222F, F295Y, and F414S and from cellubiose to gentiobiose for N222F and F414S. Using pNPG (34 mM) and hexanol 80% (vol/vol), N222F, Y295F, and F414S synthesized hexyl-ß-glycoside (HG) yields of 84.7%, 50.9%, and 54.1%, respectively, HG increased from 14.49 (TmBglA) to 22.8 mM (N222F) at 2 hr by 57.42%. However, this higher transglycosylation effect depended on that three mutants creates an environment more suited for hexanol in the active site pocket, and consequently suppressed its HG hydrolysis.


Assuntos
Glicosídeos/biossíntese , Thermotoga maritima/enzimologia , Thermotoga maritima/genética , beta-Glucosidase/genética , beta-Glucosidase/metabolismo , Alquilação , Dissacarídeos/biossíntese , Glicosídeo Hidrolases/metabolismo , Hidrólise , Microbiologia Industrial , Cinética , Engenharia Metabólica , Modelos Moleculares , Mutagênese Sítio-Dirigida , Conformação Proteica , Especificidade por Substrato
12.
J Biosci Bioeng ; 132(2): 125-131, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34078567

RESUMO

A novel ß-glucosidase was purified from pumpkin (Cucurbita moschata) seed by anion exchange chromatography and gel permeation chromatography, and its molecular mass was determined to be 42.8 kDa by gel permeation chromatography. The heterodimeric structure consisting of two subunits, free from disulfide bonds, was determined by native-PAGE analysis followed by zymography. The enzyme was maximally active at pH 4.0 and 70°C, and Vmax, Km, and kcat values were 0.078 units mg-1 protein, 2.22 mM, and 13.29 min-1, respectively, employing p-nitrophenyl-ß-d-glucopyranoside as the substrate. The high content of glycine determined by amino acid analysis implies that the enzyme possesses flexible conformations and interacts with cell membranes and walls in nature. Circular dichroism studies revealed that the high stability of the enzyme within the pH range of 2.0-10.0 is due to its reversible pH-responsive characteristics for α-helix-antiparallel ß-sheet interconversion.


Assuntos
Cucurbita , Cucurbita/metabolismo , Estabilidade Enzimática , Concentração de Íons de Hidrogênio , Cinética , Peso Molecular , Sementes/metabolismo , Especificidade por Substrato , beta-Glucosidase/metabolismo
13.
mBio ; 12(3)2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-33975944

RESUMO

Trichoderma reesei has 11 putative ß-glucosidases in its genome, playing key parts in the induction and production of cellulase. Nevertheless, the reason why the T. reesei genome encodes so many ß-glucosidases and the distinct role each ß-glucosidase plays in cellulase production remain unknown. In the present study, the cellular function and distribution of 10 known ß-glucosidases (CEL3B, CEL3E, CEL3F, CEL3H, CEL3J, CEL1A, CEL3C, CEL1B, CEL3G, and CEL3D) were explored in T. reesei, leaving out BGL1 (CEL3A), which has been well investigated. We found that the overexpression of cel3b or cel3g significantly enhanced extracellular ß-glucosidase production, whereas the overexpression of cel1b severely inhibited cellulase production by cellulose, resulting in nearly no growth of T. reesei Four types of cellular distribution patterns were observed for ß-glucosidases in T. reesei: (i) CEL3B, CEL3E, CEL3F, and CEL3G forming clearly separated protein secretion vesicles in the cytoplasm; (ii) CEL3H and CEL3J diffusing the whole endomembrane as well as the cell membrane with protein aggregation, like a reticular network; (iii) CEL1A and CEL3D in vacuoles; (iv) and CEL3C in the nucleus. ß-glucosidases CEL1A, CEL3B, CEL3E, CEL3F, CEL3G, CEL3H, and CEL3J were identified as extracellular, CEL3C and CEL3D as intracellular, and CEL1B as unknown. The extracellular ß-glucosidases CEL3B, CEL3E, CEL3F, CEL3H, and CEL3G were secreted through a tip-directed conventional secretion pathway, and CEL1A, via a vacuole-mediated pathway that was achieved without any signal peptide, while CEL3J was secreted via an unconventional protein pathway bypassing the endoplasmic reticulum (ER) and Golgi.IMPORTANCE Although ß-glucosidases play an important role in fungal cellulase induction and production, our current understanding does not provide a global perspective on ß-glucosidase function. This work comprehensively studies all the ß-glucosidases regarding their effect on cellulase production and their cellular distribution and secretion. Overexpression of cel3b or cel3g significantly enhanced ß-glucosidase production, whereas overexpression of cel1b severely inhibited cellulase production on cellulose. In addition, overexpression of cel3b, cel3e, cel3f, cel3h, cel3j, cel3c, or cel3g delayed endoglucanase (EG) production. We first identified four cellular distribution patterns of ß-glucosidases in Trichoderma reesei Specially, CEL3C was located in the nucleus. CEL3J was secreted through the nonclassical protein secretion pathway bypassing endoplasmic reticulum (ER) and Golgi. CEL1A was secreted via a vacuole-mediated conventional secretion route without a signal peptide. These findings advance our understanding of ß-glucosidase properties and secretory pathways in filamentous fungi, holding key clues for future study.


Assuntos
Proteínas Fúngicas/metabolismo , Expressão Gênica , Hypocreales/enzimologia , Hypocreales/genética , beta-Glucosidase/metabolismo , Celobiose/metabolismo , Celulase/biossíntese , Proteínas Fúngicas/genética , Regulação Fúngica da Expressão Gênica , Genes Fúngicos , Hypocreales/metabolismo , beta-Glucosidase/biossíntese , beta-Glucosidase/classificação , beta-Glucosidase/genética
14.
Int J Food Microbiol ; 350: 109242, 2021 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-34044228

RESUMO

Lactic acid fermentation is a traditional process to preserve foods and to modify their organoleptic properties. This process is generally conducted in a spontaneous way, allowing indigenous lactic acid bacteria (LAB) of the matrix and of the environment to compete and grow. The aim of this study was to better characterise LAB strains ability to modify aroma profiles in fruit and vegetable matrices, by focusing on two key enzymatic activities: ß-glucosidase and alcohol dehydrogenase (ADH). Firstly, 200 LAB isolated from Cambodian and Vietnamese fermented foods were screened for their ß-glucosidase activity and duplicate isolates identified through RAPD-PCR analysis were discarded. Thereby, 40 strains were found positive for ß-glucosidase using p-nitrophenyl-ß-D-glucopyranoside as substrate. Among them, 14 displayed an activity greater than 10 nmol/min/mg dry cell. Thirteen were identified as Lactiplantibacillus (L.) plantarum and one as L. pentosus. Secondly, four strains of different phenotypes for ß-glucosidase activity were tested for ADH activity. The highest reduction ability for hexanal and (E)-2-hexenal was obtained for Limosilactobacillus (L.) fermentum V013-1A for which no ß-glucosidase activity was detectable. The three other strains (L. plantarum C022-2B, C022-3B, and V0023-4B2) exhibited a lower reduction ability and only for hexanal. Thirdly, mashed tomatoes were fermented with these four strains individually to evaluate their ability to release volatile compounds from the tomato precursors. Fifty-eight volatile compounds were identified and quantified by HS-SPME/GC-MS. Untreated tomatoes were rich in aldehydes. The tomatoes fermented with L. plantarum strains were rich in ketones whereas those with L. fermentum were rich in alcohols. However, for the generation of terpenoids that provide flower and fruit flavours, our screening of ß-glucosidase activity was not able to explain the differences among the strains. For ADH activity, L. fermentum exhibited a high activity in fermentation as most of the target aldehydes and ketones disappeared and were replaced by their corresponding alcohols. The L. plantarum strains exhibited a lower activity but with an important substrate-selectivity diversity. A better knowledge of the functionality of each LAB strain in the food matrix will permit to predict and shape the aroma profiles of fermented food.


Assuntos
Álcool Desidrogenase/metabolismo , Alimentos e Bebidas Fermentados/microbiologia , Frutas/microbiologia , Lactobacillales/metabolismo , Verduras/microbiologia , beta-Glucosidase/metabolismo , Reatores Biológicos/microbiologia , Fermentação , Alimentos e Bebidas Fermentados/análise , Frutas/química , Cromatografia Gasosa-Espectrometria de Massas , Ácido Láctico/análise , Lactobacillales/isolamento & purificação , Odorantes/análise , Técnica de Amplificação ao Acaso de DNA Polimórfico
15.
Food Chem ; 360: 129886, 2021 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-34000634

RESUMO

This work investigated the diversity of non-Saccharomyces yeasts in main Chinese wine producing area, and discuss their potential in wine fermentation through analyzing their ß-glucosidase activity. Grapes from 44 vineyards of 9 regions were detected and a total of 395 non-Saccharomyces were identified and categorized into 16 genera, 28 species. In which, 85 non-Saccharomyces yeasts were primarily screened based on pNPG method and Bradford method, and then evaluated by ß-glucosidase environmental adaptability, substrate affinity and enzyme activity. Two selected strains were then inoculated individually or sequentially with commercial Saccharomyces cerevisiae into Gewürztraminer grape juice to detect the physiochemical indexes by HPLC and aroma compound by HS-SPME/GC-MS-FID. The results showed both non-Saccharomyces sequential inoculations increased the total aroma content, while the Candida glabrata strain D18 significantly increased the typicality and complexity of the floral and fruity aroma in Gewürztraminer wines, demonstrated its potential in wine fermentation.


Assuntos
Saccharomyces cerevisiae/metabolismo , Vitis/metabolismo , Vinho/análise , Leveduras/metabolismo , beta-Glucosidase/metabolismo , China , Fermentação , Odorantes/análise , Vitis/química , Leveduras/química
16.
Methods Mol Biol ; 2290: 203-214, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34009592

RESUMO

Vegetable oil-derived biodiesels have a major quality problem due to the presence of precipitates formed by steryl glucosides, which clog filters and injectors of diesel engines. An efficient, scalable, and cost-effective method to hydrolyze steryl glucosides using thermostable enzymes has been developed. Here, methods to discover, express in recombinant microorganisms and manufacture enzymes with SGase activity, as well as methods to treat biodiesel with such enzymes, and to measure the content of steryl glucosides in biodiesel samples are presented.


Assuntos
Glucosídeos/química , Fitosteróis/química , beta-Glucosidase/metabolismo , Biocombustíveis/análise , Clonagem Molecular/métodos , Enzimas/química , Hidrólise , Óleos Vegetais , beta-Glucosidase/biossíntese
17.
Int J Mol Sci ; 22(7)2021 Mar 29.
Artigo em Inglês | MEDLINE | ID: mdl-33805379

RESUMO

ß-Glucosidase is a microbial cellulose multienzyme that plays an important role in the regulation of the entire cellulose hydrolysis process, which is the rate-limiting step in bacterial carbon cycling in marine environments. Despite its importance in coral reefs, the diversity of ß-glucosidase-producing bacteria, their genes, and enzymatic characteristics are poorly understood. In this study, 87 ß-glucosidase-producing cultivable bacteria were screened from 6 genera of corals. The isolates were assigned to 21 genera, distributed among three groups: Proteobacteria, Firmicutes, and Actinobacteria. In addition, metagenomics was used to explore the genetic diversity of bacterial ß-glucosidase enzymes associated with scleractinian corals, which revealed that these enzymes mainly belong to the glycosidase hydrolase family 3 (GH3). Finally, a novel recombinant ß-glucosidase, referred to as Mg9373, encompassing 670 amino acids and a molecular mass of 75.2 kDa, was classified as a member of the GH3 family and successfully expressed and characterized. Mg9373 exhibited excellent tolerance to ethanol, NaCl, and glucose. Collectively, these results suggest that the diversity of ß-glucosidase-producing bacteria and genes associated with scleractinian corals is high and novel, indicating great potential for applications in the food industry and agriculture.


Assuntos
Antozoários/microbiologia , Bactérias/enzimologia , Microbiota/genética , beta-Glucosidase/genética , Animais , Bactérias/genética , Metagenômica , Filogenia , beta-Glucosidase/metabolismo
18.
Colloids Surf B Biointerfaces ; 203: 111761, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33872829

RESUMO

In this study, surficial interactions of glutaraldehyde (GA) as an important crosslinker agent with the ß-glucosidase (BGL) enzyme surface were investigated by theoretical methods. Since the inherent constraints of experimental methods limit their application to find the molecular perspective of these significant interactions in enzyme immobilization, theoretical methods were used as a complementary tool to understand this concept. The Minnesota density functional calculations showed that the chair conformations of the oxane-2,6-diol form of the GA were more stable than its free aldehyde form. MD simulations of propylamine-GA molecules, as a representative of attached-GA, in aqueous solutions of different concentrations were done to determine the molecular basis of surficial interactions with the BGL surface. The root mean square fluctuation (RMSF) demonstrated that the maximum flexibility of the BGL enzyme belonged to 460-480 residues in all solutions. Based on the spatial distribution function (SDF) analysis, the active site entrance was the most favored region to accumulate solute molecules. Radial distribution function (RDF) results showed that all forms of propylamine-GA molecules interacted from their head side with the lysine residues of BGL, which Lys247, Lys376, and Lys384 were found to be the most interactive lysine residues. Also, hydrogen bond (HB) analysis from two viewpoints confirmed HB formation possibility between propylamine-GA molecules and these lysine residues. These results explained which regions of the BGL have the maximum possibility to interact and link to GA and help us in understanding the process of enzyme immobilization.


Assuntos
Simulação de Dinâmica Molecular , beta-Glucosidase , Domínio Catalítico , Ligação de Hidrogênio , Água , beta-Glucosidase/metabolismo
19.
J Food Sci ; 86(6): 2327-2345, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33929752

RESUMO

ß-glucosidase is a pivotal enzyme that hydrolyzes bound volatile aromatic compounds. However, the activity of ß-glucosidase in winemaking and the mechanism by which it affects the flavor and taste of wines have not been fully investigated. In this study, we profiled the characteristics of ß-glucosidase derived from wine-related yeasts isolated from different wine-making regions in China, and analyzed the enzyme activity from different parts of the cells under aerobic and anaerobic conditions. A total of 56 strains of wine-related yeasts producing ß-glucosidases were screened using the YNB-C medium (YNB 6.7 g L-1 , cellobiose 5 g L-1 , pH 5.0). We found that strain Clavispora lusitaniae C117 produced the highest enzyme activity (152.39 µmol pNP ml-1 h-1 ). In most strains, ß-glucosidase were located in whole cells (periplasmic space) and permeabilized cells (intracellular). The non-Saccharomyces species had the highest enzymatic activity in a strain-dependent manner. Under aerobic conditions, C. lusitaniae C117, Hanseniaspora guilliermondii A27-3-4, Metschnikowia pulcherrima F-1-6, and Pichia anomala C84 had the highest ß-glucosidase activity. We further investigated the ß-glucosidase activity during the wine fermentation and the effects of sugar, pH, temperature, and ethanol on the enzyme activities of P. anomala C84 and commercial Saccharomyces yeast strains RC212 and VL1. The presence of fructose, glucose, and sucrose strongly inhibited enzyme activity. Similarly, low pH and low temperature inhibited the activity of ß-glucosidase, whereas ethanol promoted enzyme activity. Our findings provide a theoretical basis on understanding the different yeast characteristics of ß-glucosidase and their potential application for further improving wine aroma complexity.


Assuntos
Hanseniaspora/enzimologia , Metschnikowia/enzimologia , Odorantes/análise , Saccharomycetales/enzimologia , Vinho/análise , beta-Glucosidase/metabolismo
20.
Molecules ; 26(7)2021 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-33808362

RESUMO

Olive mill wastewater (OMW) contains valuable and interesting bioactive compounds, among which is hydroxytyrosol, which is characterized by a remarkable antioxidant activity. Due to the health claims related to olive polyphenols, the aim of this study was to obtain an extract from OMW with an increased level of hydroxytyrosol by means of microbial enzymatic activity. For this purpose, four commercial adsorbent resins were selected and tested. The beta-glucosidase and esterase activity of strains of Wickerhamomyces anomalus, Lactiplantibacillus plantarum, and Saccharomyces cerevisiae were also investigated and compared to those of a commercial enzyme and an Aspergillus niger strain. The W. anomalus strain showed the best enzymatic performances. The SP207 resin showed the best efficiency in selective recovery of hydroxytyrosol, tyrosol, oleuropein, and total phenols. The bioconversion test of the OMW extract was assessed by using both culture broths and pellets of the tested strains. The results demonstrated that the pellets of W. anomalus and L. plantarum were the most effective in hydroxytyrosol increasing in phenolic extract. The interesting results suggest the possibility to study new formulations of OMW phenolic extracts with multifunctional microorganisms.


Assuntos
Fungos/metabolismo , Olea/química , Fenóis/química , Extratos Vegetais/química , Eliminação de Resíduos Líquidos , beta-Glucosidase/metabolismo , Lactobacillaceae/metabolismo , Probióticos
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