Functional characterization and target discovery of glycoside hydrolases from the digestome of the lower termite Coptotermes gestroi

Cairo, João Paulo L. Franco; Leonardo, Flávia C.; Alvarez, Thabata M.; Ribeiro, Daniel; Büchli, Fernanda; Costa–Leonardo, Ana Maria; Carazzolle, Marcelo Falsarella; Costa, Fernando Ferreira; Leme, Adriana Franco Paes; Pereira, Gonçalo Amarante Guimarães; Squina, Fábio M.

doi:10.1186/1754-6834-4-50

Cited by 38 publications

(30 citation statements)

References 53 publications

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“…The kinetic parameters were calculated from initial velocities using p-NPG at 0.5-14 mM concentrations. Reactions were carried out in 50 mM sodium acetate buffer (pH 5.5) with 0.5 μg purified BGs at 40°C and was stopped by adding 1 M Na 2 CO 3 [24]. The rate of pnitrophenol (p-NP) released was measured every 1 min by absorbance at 410 nm.…”

Section: Kinetic Parameter Determination and Mutant Activity Assaymentioning

confidence: 99%

Identification and Characterization of Two Endogenous β-Glucosidases from the Termite Coptotermes formosanus

Feng

Sun

et al. 2015

Appl Biochem Biotechnol

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Coptotermes formosanus is a well-known wood-feeding termite that can degrade lignocellulose polysaccharides efficiently with its unique multi-enzyme catalysis system. β-glucosidase (BG) is one of the important cellulases in its enzyme system. However, there may present multiple endogenous BGs in termite digestive system for various properties and functions. This study aims to characterize two BG homologs and reveal their potential coordinative effect. In this study, two endogenous BG homologs (CfGlu1B and CfGlu1C) from C. formosanus showed different substrate specificity. CfGlu1B favors cellobiose while CfGlu1C favors sucrose. Besides, CfGlu1C exhibited higher alkali resistance than CfGlu1B. Kinetic analysis revealed that CfGlu1B enzyme's activity toward p-NP-β-D-glucopyranoside (p-NPG) was higher than that of CfGlu1C, and the difference mainly attributes to the turnover number (K cat). In addition, the activity assay showed significant synergistic action of CfGlu1B and CfGlu1C in degrading D-lactosum. For effect of metals, Cu(2+) inhibited both enzymes and Ca(2+) increased the activity of CfGlu1C but not CfGlu1B. Site-directed mutagenesis analysis indicated that both enzymes lost activities when residues E190 of CfGlu1B and E168 of CfGlu1C were mutated to alanine, respectively, which were essential active centers of the GHF1 enzymes. Moreover, mutation H252N increased the activity of enzyme CfGlu1C by 2.1-fold. This study implies interesting possibilities for better practical biotechnological use in green energy production.

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Section: Kinetic Parameter Determination and Mutant Activity Assaymentioning

confidence: 99%

Identification and Characterization of Two Endogenous β-Glucosidases from the Termite Coptotermes formosanus

Feng

Sun

et al. 2015

Appl Biochem Biotechnol

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“…In the termite gut, carbohydrate-active enzymes (CAZymes), such as cellulases and hemicellulases, are secreted (i.e., both symbiotic and endogenous enzymes). Additionally, a set of pro-oxidant, antioxidant, and detoxification enzymes (PADs) is also present [5–7]. Among the PADs found in termites, superoxide dismutase (SOD), catalases (CATs), glutathione S-transferase (GST), and aldo–keto reductases (AKRs) have been studied in detail because the transcription of mRNAs encoding these enzymes is upregulated in response to lignocellulose degradation [8].…”

Section: Introductionmentioning

confidence: 99%

The Coptotermes gestroi aldo–keto reductase: a multipurpose enzyme for biorefinery applications

Tramontina

Cairo

Liberato

et al. 2017

Biotechnol Biofuels

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BackgroundIn nature, termites can be considered as a model biological system for biofuel research based on their remarkable efficiency for lignocellulosic biomass conversion. Redox enzymes are of interest in second-generation ethanol production because they promote synergic enzymatic activity with classical hydrolases for lignocellulose saccharification and inactivate fermentation inhibitory compounds produced after lignocellulose pretreatment steps.ResultsIn the present study, the biochemical and structural characteristics of the Coptotermes gestroi aldo–keto reductase (CgAKR-1) were comprehensively investigated. CgAKR-1 displayed major structural differences compared with others AKRs, including the differences in the amino acid composition of the substrate-binding site, providing basis for classification as a founding member of a new AKR subfamily (family AKR1 I). Immunolocalization assays with anti-CgAKR-1 antibodies resulted in strong fluorescence in the salivary gland, proventriculus, and foregut. CgAKR-1 supplementation caused a 32% reduction in phenolic aldehydes, such as furfural, which act as fermentation inhibitors of hemicellulosic hydrolysates, and improved ethanol fermentation by the xylose-fermenting yeast Scheffersomyces stipitis by 45%. We observed synergistic enzymatic interactions between CgAKR-1 and commercial cellulosic cocktail for sugarcane bagasse saccharification, with a maximum synergism degree of 2.17 for sugar release. Our data indicated that additive enzymatic activity could be mediated by reactive oxygen species because CgAKR-1 could produce hydrogen peroxide.ConclusionIn summary, we identified the founding member of an AKRI subfamily with a potential role in the termite digestome. CgAKR-1 was found to be a multipurpose enzyme with potential biotechnological applications. The present work provided a basis for the development and application of integrative and multipurpose enzymes in the bioethanol production chain.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0688-6) contains supplementary material, which is available to authorized users.

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“…Biomass-degrading microorganisms also often act in synergy in order to optimize their growth efforts by minimizing the energetic expense for production of the required degradation enzymes and by maximizing benefits of their common action. For these reasons, metagenomic studies focusing on whole microbial communities rather than isolated organisms, are more likely to identify interesting enzymes for lignocellulose degradation, especially when these communities are derived from specialized communities from lignocellulose-rich ecological niches, such as those from soil, or from specific natural or artificial environments such as compost, or the digestive tract of large herbivores or termites [4–7]. …”

Section: Introductionmentioning

confidence: 99%

Composting-Like Conditions Are More Efficient for Enrichment and Diversity of Organisms Containing Cellulase-Encoding Genes than Submerged Cultures

et al. 2016

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Cost-effective biofuel production from lignocellulosic biomass depends on efficient degradation of the plant cell wall. One of the major obstacles for the development of a cost-efficient process is the lack of resistance of currently used fungal enzymes to harsh conditions such as high temperature. Adapted, thermophilic microbial communities provide a huge reservoir of potentially interesting lignocellulose-degrading enzymes for improvement of the cellulose hydrolysis step. In order to identify such enzymes, a leaf and wood chip compost was enriched on a mixture of thermo-chemically pretreated wheat straw, poplar and Miscanthus under thermophile conditions, but in two different set-ups. Unexpectedly, metagenome sequencing revealed that incubation of the lignocellulosic substrate with compost as inoculum in a suspension culture resulted in an impoverishment of putative cellulase- and hemicellulase-encoding genes. However, mimicking composting conditions without liquid phase yielded a high number and diversity of glycoside hydrolase genes and an enrichment of genes encoding cellulose binding domains. These identified genes were most closely related to species from Actinobacteria, which seem to constitute important players of lignocellulose degradation under the applied conditions. The study highlights that subtle changes in an enrichment set-up can have an important impact on composition and functions of the microcosm. Composting-like conditions were found to be the most successful method for enrichment in species with high biomass degrading capacity.

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Functional characterization and target discovery of glycoside hydrolases from the digestome of the lower termite Coptotermes gestroi

Cited by 38 publications

References 53 publications

Identification and Characterization of Two Endogenous β-Glucosidases from the Termite Coptotermes formosanus

Identification and Characterization of Two Endogenous β-Glucosidases from the Termite Coptotermes formosanus

The Coptotermes gestroi aldo–keto reductase: a multipurpose enzyme for biorefinery applications

Composting-Like Conditions Are More Efficient for Enrichment and Diversity of Organisms Containing Cellulase-Encoding Genes than Submerged Cultures

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