Synergic activity of Cel8Pa β-1,4 endoglucanase and Bg1Pa β-glucosidase from Paenibacillus xylanivorans A59 in beta-glucan conversion

Ghio, Silvina; Bradanini, María B.; Garrido, Mercedes María; Ontañon, Ornella Mailén; Piccinni, Florencia Elizabeth; Villegas, Rubén Marrero Díaz de; Talia, Paola; Campos, Eleonora

doi:10.1016/j.btre.2020.e00526

Cited by 14 publications

(11 citation statements)

References 40 publications

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“…In addition, there are also endoglucanases that have a wider substrate specificity and are active on hemicelluloses like xyloglucan, xylan and arabinoxylan such as the endoglucanase family Cel7, which could explain the low amount of xylose released by EndoB (Vlasenko et al 2010;Hua et al 2018). The levels of the oligosaccharides were higher for the formulations, except for EndoC, which hints to synergistical action of the endoglucanase enzymes with other components of the enzyme formulations like β-glucosidases as reported in previous studies (Song et al 2016;Ghio et al 2020). On the short fiber hardwood pulp, the purified endoglucanases showed a similar pattern as for the long fiber softwood pulp, but a much lower amount of the different sugars was released (Fig.…”

Section: Resultsmentioning

confidence: 54%

Mechanistic investigation of the effect of endoglucanases related to pulp refining

et al. 2022

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Endoglucanases are increasingly being touted as the ultimate solution for reducing energy consumption during the refining process in the pulp and paper industry. However, due to the high variety of endoglucanases in different enzyme formulations, these perform heterogeneously when applied to different pulps. In this study, the effect of four endoglucanases on softwood and hardwood pulp was studied using confocal laser scanning microscopy (CLSM) after addition of fluorescently labelled carbohydrate binding modules (CBMs). Nuclear magnetic resonance (NMR) analysis and high-performance liquid chromatography quantification of released oligo- and monosaccharides was performed for in-depth mechanistical investigation. Changes in the crystallinity levels caused by enzymatic degradation of amorphous regions were monitored by incubation with two different CBMs from Caldicellulosiruptor bescii and from Thermobifida fusca with high preference to either amorphous or crystalline regions of cellulose, respectively. When dosed at identical activity on the endoglucanase specific CellG5 substrate, CLSM analysis indicated the highest decrease of amorphous regions for those endoglucanases which were also most active in laboratory refining trials and which released highest amounts of cellooligomers from pulp. Using 13C-NMR analysis, an increase in para-crystalline cellulose caused by enzyme application was observed. Release of reducing sugars was determined at identical CellG5 dosage, indicating a high variance between the enzymes, especially when complex enzyme formulations were used. Scanning electron microscopy images were obtained for visualization of the endoglucanase activity. The results of mechanistical studies indicate that reduction of amorphous moieties of pulp by endoglucanases is especially beneficial for the refining process.

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Section: Resultsmentioning

confidence: 54%

Mechanistic investigation of the effect of endoglucanases related to pulp refining

et al. 2022

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“…Moreover, the suppression of its endo-β-1,4-glucanase activity exerted by 2.0 M NaCl was only observed to be approximately 5%, implying that it was a relatively salt-tolerant enzyme. These findings suggested that the salt tolerance of rGluL was superior to that of Paenibacillus xylanivorans A59 GH8 endo-β-1,4-glucanase ( Ghio et al, 2020 ), which showed biocatalytic activity below 50% in the presence of NaCl between 0.5 and 2.0 M.…”

Section: Resultsmentioning

confidence: 99%

Novel, acidic, and cold-adapted glycoside hydrolase family 8 endo-β-1,4-glucanase from an Antarctic lichen-associated bacterium, Lichenicola cladoniae PAMC 26568

Kim

Kim²,

Lee³

et al. 2022

Front. Microbiol.

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Endo-β-1,4-glucanase is a crucial glycoside hydrolase (GH) involved in the decomposition of cellulosic materials. In this study, to discover a novel cold-adapted β-1,4-D-glucan-degrading enzyme, the gene coding for an extracellular endo-β-1,4-glucanase (GluL) from Lichenicola cladoniae PAMC 26568, an Antarctic lichen (Cladonia borealis)-associated bacterium, was identified and recombinantly expressed in Escherichia coli BL21. The GluL gene (1044-bp) encoded a non-modular polypeptide consisting of a single catalytic GH8 domain, which shared the highest sequence identity of 55% with that of an uncharacterized protein from Gluconacetobacter takamatsuzukensis (WP_182950054). The recombinant endo-β-1,4-glucanase (rGluL: 38.0 kDa) most efficiently degraded sodium carboxymethylcellulose (CMC) at pH 4.0 and 45°C, and showed approximately 23% of its maximum degradation activity even at 3°C. The biocatalytic activity of rGluL was noticeably enhanced by >1.3-fold in the presence of 1 mM Mn2+ or NaCl at concentrations between 0.1 and 0.5 M, whereas the enzyme was considerably downregulated by 1 mM Hg2+ and Fe2+ together with 5 mM N-bromosuccinimide and 0.5% sodium dodecyl sulfate. rGluL is a true endo-β-1,4-glucanase, which could preferentially decompose D-cellooligosaccharides consisting of 3 to 6 D-glucose, CMC, and barley β-glucan, without other additional glycoside hydrolase activities. The specific activity (15.1 U mg–1) and kcat/Km value (6.35 mg–1 s–1mL) of rGluL toward barley β-glucan were approximately 1.8- and 2.2-fold higher, respectively, compared to its specific activity (8.3 U mg–1) and kcat/Km value (2.83 mg–1 s–1mL) toward CMC. The enzymatic hydrolysis of CMC, D-cellotetraose, and D-cellohexaose yielded primarily D-cellobiose, accompanied by D-glucose, D-cellotriose, and D-cellotetraose. However, the cleavage of D-cellopentaose by rGluL resulted in the production of only D-cellobiose and D-cellotriose. The findings of the present study imply that rGluL is a novel, acidic, and cold-adapted GH8 endo-β-1,4-glucanase with high specific activity, which can be exploited as a promising candidate in low-temperature processes including textile and food processes.

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“…Figure A shows that the glucose concentration increased at the start but decreased after 12 h, whereas the cellobiose concentration slightly increased at the start but decreased after 8 h. The cellulase enzymes include endoglucanase, exoglucanase (cellobiohydrolase), and β-glucosidase, which act synergistically. The endoglucanase generates reducing and nonreducing chain ends for cellobiohydrolase that releases cellobiose (dimer), which is converted to glucose by β-glucosidase . As Cellic CTec2 has high cellulase and β-glucosidase activities, it produces an elevated level of glucose.…”

Section: Resultsmentioning

confidence: 99%

“…The endoglucanase generates reducing and nonreducing chain ends for cellobiohydrolase that releases cellobiose (dimer), which is converted to glucose by β-glucosidase. 45 As Cellic CTec2 has high cellulase and β-glucosidase activities, it produces an elevated level of glucose. Cellic CTec2 was reported for efficient hydrolysis of hydrothermally pretreated wheat straw.…”

Section: Fermentation Kinetics (Flask)mentioning

confidence: 99%

Cellulase Addition and Pre-hydrolysis Effect of High Solid Fed-Batch Simultaneous Saccharification and Ethanol Fermentation from a Combined Pretreated Oil Palm Trunk

et al. 2021

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In the current study, alkaline hydrogen peroxide pretreated oil palm trunk fibers were subjected to ethanol production via simultaneous saccharification and fermentation (SSF). The effect of high substrate loading, enzyme and substrate feeding strategy, and influence of a pre-hydrolysis step in SSF was studied to scale up ethanol production. In the enzyme feeding strategy, the addition of an enzyme at the start of fed-batch SSF significantly (p < 0.05) increased ethanol concentration to 51.05 g/L, ethanol productivity (Q P ) to 0.61 g/L•h, and ethanol yield (Y P/S ) to 0.31 g/g, with a theoretical ethanol yield of 60.65%. Furthermore, the initial velocity of the enzyme (V 0 ) in the first 8 h was 2.27 (g/h) with a glucose concentration of 18.17 g/L. On the other hand, the substrate feeding strategy and pre-hydrolysis simultaneous saccharification and fermentation (PSSF) process were studied in a 1 L fermenter. PSSF in fed batch with 10 and 20% (w/v) significantly improved enzyme hydrolysis, circumvent the problems of high viscosity, reduced overall fermentation time, and gave the highest ethanol concentration of 51.66 g/L, ethanol productivity (Q P ) of 0.72 g/L•h, ethanol yield (Y P/S ) of 0.31 g/g, and theoretical ethanol yield of 60.66%. In addition, PSSF with 10 and 20% significantly increased the initial velocity of the enzyme (V 0 ) to 4.64 and 4.40 (g/h) and glucose concentration to 37.14 and 35.27 g/L, respectively. This result indicated that ethanol production by PSSF along with substrate feeding could enhance ethanol production efficiently.

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Synergic activity of Cel8Pa β-1,4 endoglucanase and Bg1Pa β-glucosidase from Paenibacillus xylanivorans A59 in beta-glucan conversion

Cited by 14 publications

References 40 publications

Mechanistic investigation of the effect of endoglucanases related to pulp refining

Mechanistic investigation of the effect of endoglucanases related to pulp refining

Novel, acidic, and cold-adapted glycoside hydrolase family 8 endo-β-1,4-glucanase from an Antarctic lichen-associated bacterium, Lichenicola cladoniae PAMC 26568

Cellulase Addition and Pre-hydrolysis Effect of High Solid Fed-Batch Simultaneous Saccharification and Ethanol Fermentation from a Combined Pretreated Oil Palm Trunk

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