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Updated: Jun 25, 2025
Author Spotlight: Integrating Biochemical Functions of β-Glucanases and Peroxidase Enzymes in Wheat-RWA Interaction
Published on: July 26, 2024
Expression and characterization of a novel microbial GH9 glucanase, IDSGLUC9-4, isolated from sheep rumen
Yongzhen Zhu1,2,3, Shuning Bai2,3, Nuo Li2,3
1Jilin Key Laboratory for Molecular Biology of Special Economic Animals, Institute of Special Animal and Plant Sciences, Chinese Academy of Agricultural Sciences, Changchun 130112, China.
Objective:
This study aimed to identify and characterize a novel endo-β-glucanase, IDSGLUC9-4, from the rumen metatranscriptome of Hu sheep.
Methods:
A novel endo-β-glucanase, IDSGLUC9-4, was heterologously expressed in Escherichia coli and biochemically characterized. The optimal temperature and pH of recombinant IDSGLUC9-4 were determined. Subsequently, substrate specificity of the enzyme was assessed using mixed-linked glucans including barley β-glucan and Icelandic moss lichenan. Thin-layer chromatography (TLC), high-performance liquid chromatography (HPLC), matrix assisted laser desorption ionization time of flight mass spectrometry analyses were conducted to determine the products released from polysaccharides and cello-oligosaccharides substrates.
Results:
The recombinant IDSGLUC9-4 exhibited temperature and pH optima of 40°C and pH 6.0, respectively. It exclusively hydrolyzed mixed-linked glucans, with significant activity observed for barley β-glucan (109.59±3.61 μmol/mg min) and Icelandic moss lichenan (35.35±1.55 μmol/mg min). TLC and HPLC analyses revealed that IDSGLUC9-4 primarily released cellobiose, cellotriose, and cellotetraose from polysaccharide substrates. Furthermore, after 48 h of reaction, IDSGLUC9-4 removed most of the glucose, indicating transglycosylation activity alongside its endo-glucanase activity.
Conclusion:
The recombinant IDSGLUC9-4 was a relatively acid-resistant, mesophilic endo-glucanase (EC 3.2.1.4) that hydrolyzed glucan-like substrates, generating predominantly G3 and G4 oligosaccharides, and which appeared to have glycosylation activity. These findings provided insights into the substrate specificity and product profiles of rumen-derived GH9 glucanases and contributed to the expanding knowledge of cellulolytic enzymes and novel herbivore rumen enzymes in general.
