Dataset Information

Evolution and functional characterization of CAZymes belonging to subfamily 10 of glycoside hydrolase family 5 (GH5_10) in two species of phytophagous beetles.

ABSTRACT: Hemicelluloses, such as xyloglucan, xylan and mannans, consist of a heterogeneous array of plant-derived polysaccharides that form the plant cell wall. These polysaccharides differ from each other in their structure and physiochemical properties, but they share a ?-(1,4)-linked sugar backbone. Hemicelluloses can be hydrolyzed by plant-cell-wall-degrading enzymes (PCWDEs), which are widely distributed in phytopathogenic microbes. Recently, it has become apparent that phytophagous beetles also produce their own PCWDEs. Our previous work identified genes encoding putative mannanases belonging to the subfamily 10 of glycoside hydrolase (GH) family 5 (GH5_10) in the genomes of the leaf beetle, Gastrophysa viridula (Chrysomelidae, Chrysomelinae; one gene), and of the bean beetle, Callosobruchus maculatus (Chrysomelidae, Bruchinae; four genes). In contrast to proteins from other GH5 subfamilies, GH5_10 proteins are patchily distributed within the tree of life and have so far hardly been investigated. We addressed the following questions: Are beetle-derived GH5_10s active PCWDEs? How did they evolve? What is their physiological function? Using heterologous protein expression and enzymatic assays, we show that the G. viridula GH5_10 protein is an endo-?-1,4-mannanase. We also demonstrate that only one out of four C. maculatus GH5_10 proteins is an endo-?-1,4-mannanase, which has additional activity on carboxymethyl cellulose. Unexpectedly, another C. maculatus GH5_10 protein has evolved to use xylan instead of mannans as a substrate. RNAi experiments in G. viridula indicate (i) that the sole GH5_10 protein is responsible for breaking down mannans in the gut and (ii) that this breakdown may rather be accessory and may facilitate access to plant cell content, which is rich in nitrogen and simple sugars. Phylogenetic analyses indicate that coleopteran-derived GH5_10 proteins cluster together with Chelicerata-derived ones. Interestingly, other insect-derived GH5_10 proteins cluster elsewhere, suggesting insects have several independent evolutionary origins.

SUBMITTER: Busch A

PROVIDER: S-EPMC5576741 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Evolution and functional characterization of CAZymes belonging to subfamily 10 of glycoside hydrolase family 5 (GH5_10) in two species of phytophagous beetles.

Busch André A Kunert Grit G Heckel David G DG Pauchet Yannick Y

PloS one 20170830 8

Hemicelluloses, such as xyloglucan, xylan and mannans, consist of a heterogeneous array of plant-derived polysaccharides that form the plant cell wall. These polysaccharides differ from each other in their structure and physiochemical properties, but they share a β-(1,4)-linked sugar backbone. Hemicelluloses can be hydrolyzed by plant-cell-wall-degrading enzymes (PCWDEs), which are widely distributed in phytopathogenic microbes. Recently, it has become apparent that phytophagous beetles also pro ...[more]

PMID: 28854242

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Project description:As classified by the Carbohydrate-Active Enzymes (CAZy) database, enzymes in glycoside hydrolase (GH) family 10 (GH10) are all monospecific or bifunctional xylanases (except a tomatinase), and no endo-β-1,4-glucanase has been reported in the family. Here, we identified Arcticibacterium luteifluviistationis carboxymethyl cellulase (AlCMCase) as a GH10 endo-β-1,4-glucanase. AlCMCase originated from an Arctic marine bacterium, Arcticibacterium luteifluviistationis SM1504T It shows low identity (<35%) with other GH10 xylanases. The gene encoding AlCMCase was overexpressed in Escherichia coli Biochemical characterization showed that recombinant AlCMCase is a cold-adapted and salt-tolerant enzyme. AlCMCase hydrolyzes cello- and xylo-configured substrates via an endoaction mode. However, in comparison to its significant cellulase activity, the xylanase activity of AlCMCase is negligible. Correspondingly, AlCMCase has remarkable binding capacity for cello-oligosaccharides but no obvious binding capacity for xylo-oligosaccharides. AlCMCase and its homologs are grouped into a branch separate from other GH10 xylanases in a phylogenetic tree, and two homologs also displayed the same substrate specificity as AlCMCase. These results suggest that AlCMCase and its homologs form a novel subfamily of GH10 enzymes that have robust endo-β-1,4-glucanase activity. In addition, given the cold-adapted and salt-tolerant characters of AlCMCase, it may be a candidate biocatalyst under certain industrial conditions, such as low temperature or high salinity.IMPORTANCE Cellulase and xylanase have been widely used in the textile, pulp and paper, animal feed, and food-processing industries. Exploring novel cellulases and xylanases for biocatalysts continues to be a hot issue. Enzymes derived from the polar seas might have novel hydrolysis patterns, substrate specificities, or extremophilic properties that have great potential for both fundamental research and industrial applications. Here, we identified a novel cold-adapted and salt-tolerant endo-β-1,4-glucanase, AlCMCase, from an Arctic marine bacterium. It may be useful in certain industrial processes, such as under low temperature or high salinity. Moreover, AlCMCase is a bifunctional representative of glycoside hydrolase (GH) family 10 that preferentially hydrolyzes β-1,4-glucans. With its homologs, it represents a new subfamily in this family. Thus, this study sheds new light on the substrate specificity of GH10.

Dataset Information

Evolution and functional characterization of CAZymes belonging to subfamily 10 of glycoside hydrolase family 5 (GH5_10) in two species of phytophagous beetles.

Publications

Evolution and functional characterization of CAZymes belonging to subfamily 10 of glycoside hydrolase family 5 (GH5_10) in two species of phytophagous beetles.

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