Journal
PREPARATIVE BIOCHEMISTRY & BIOTECHNOLOGY
Volume 51, Issue 9, Pages 881-891Publisher
TAYLOR & FRANCIS INC
DOI: 10.1080/10826068.2020.1870235
Keywords
Alkaline; cold-active; mannanase; mannans; Verrucomicrobiae
Categories
Funding
- Hubei Provincial Natural Science Foundation of China [2020CFB803]
- China Ocean Mineral Resources Research and Development Association [DY135-B2-06]
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A new beta-mannanase, ManDG1235 from Verrucomicrobiae DG1235, was characterized biochemically and its enzymatic properties were revealed. The purified rManDG1235 exhibited cold-active enzyme properties, with optimal activity at 20 degrees Celsius and high activity at pH 8. A structural model suggested that ManDG1235 has four mannose-binding subsites and an exo-acting feature for specifically cleaving mannobiose from the non-reducing end of the substrate.
Mannanases catalyze the cleavage of beta-1,4-mannosidic linkages in mannans and have various applications in different biotechnological industries. In this study, a new beta-mannanase from Verrucomicrobiae DG1235 (ManDG1235) was biochemically characterized and its enzymatic properties were revealed. Amino acid alignment indicated that ManDG1235 belonged to glycoside hydrolase family 26 and shared a low amino acid sequence identity to reported beta-mannanases (up to 50% for CjMan26C from Cellvibrio japonicus). ManDG1235 was expressed in Escherichia coli. Purified ManDG1235 (rManDG1235) exhibited the typical properties of cold-active enzymes, including high activity at low temperature (optimal at 20 degrees C) and thermal instability. The maximum activity of rManDG1235 was achieved at pH 8, suggesting that it is a mildly alkaline beta-mannanase. rManDG1235 was able to hydrolyze a variety of mannan substrates and was active toward certain types of glucans. A structural model that was built by homology modeling suggested that ManDG1235 had four mannose-binding subsites which were symmetrically arranged in the active-site cleft. A long loop linking beta 2 and alpha 2 as in CjMan26C creates a steric border in the glycone region of active-site cleft which probably leads to the exo-acting feature of ManDG1235, for specifically cleaving mannobiose from the non-reducing end of the substrate.
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