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| Identification and structural analysis of an alkali-tolerant cellulase from Bacillus licheniformis |
| Received:April 02, 2025 |
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| KeyWord:Bacillus licheniformis;protein structure;cellulase;enzymatic property;GH9 family |
| Author Name | Affiliation | E-mail | | FENG Yichang | College of Agriculture, Ningxia University, Yinchuan 750021, China | | | KONG Xinru | College of Agriculture, Ningxia University, Yinchuan 750021, China | | | JIANG Xin | College of Agriculture, Ningxia University, Yinchuan 750021, China | | | LI Yulong | College of Agriculture, Ningxia University, Yinchuan 750021, China | yulongli@nxu.edu.cn |
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| Abstract: |
| In search of efficient and powerful lignocellulose-degrading enzymes, a cellulase-producing Bacillus licheniformis strain YC36 was isolated from agricultural straw compost samples. The cellulase gene was cloned via homologous cloning techniques, inserted into the pET-32a（+）expression vector, and heterologously expressed in Escherichia coli BL21（DE3）. The recombinant protein was purified using affinity chromatography, and its enzymatic properties were systematically characterized. Bioinformatic approaches were also employed to elucidate its alkali tolerance mechanism. The results revealed that the molecular weight of the recombinant cellulase from YC36 was 70.4 kDa. Enzymatic assays demonstrated that the optimal reaction conditions were pH 9.0 and 55 ℃. The enzyme retained over 80% of its activity within a pH range of 5.0-9.0 and a temperature range of 50-65 ℃. Thermostability analysis showed that the enzyme maintained 85% activity after incubation at 55 ℃ for 60 minutes. Regarding ionic effects, Mg2+ significantly enhanced enzymatic activity, while Cu2+ exerted a strong inhibitory effect. Bioinformatic analysis indicated that YC36 cellulase possesses a GH9-CBM3 domain structure, a highly hydrophobic core, and alkaline-active site residues, which collectively confer excellent thermostability and broad pH adaptability. The study investigated the functional characteristics of YC-36′s GH9 cellulase under thermophilic and alkaline conditions. Through analysis of its critical structural features including the GH9-CBM3 domain, highly hydrophobic core, and alkaline-active site, we elucidated the molecular mechanisms underlying its thermostability and pH adaptability. These findings establish a theoretical foundation for developing industrial enzyme preparations with superior environmental adaptability while providing valuable genetic resources for enzymatic engineering applications. |
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