Structure and sequence analysis-based engineering of pullulanase from Anoxybacillus sp LM18-11 for improved thermostability
文献类型：期刊论文
| 作者 | Li, Shu-fang; Xu, Jian-yong; Bao, Yun-juan; Zheng, Hong-chen; Song, Hui |
| 刊名 | JOURNAL OF BIOTECHNOLOGY
 |
| 出版日期 | 2015-09-20 |
| 卷号 | 210页码:8-14 |
| 关键词 | Pullulanase Thermostability Consensus method Flexible region Anoxybacillus sp LM18-11 |
| 英文摘要 | Pullulanase (EC 3.2.1.41) is a well-known starch-debranching enzyme. Enhancing the thermostability of Pullulanase is required for industrial application. In this study, we used two methods to improve the thermostability of the pullulanase from Anoxybacillus sp. LM18-11; these methods were the modified amino acid consensus method combined with the analyses of the residue water-exposed surface (ACC) and the deletion of flexible domains. Four mutants (Y477A, Y175C, L215C and R473E) were obtained via the modified consensus method exhibited varying degrees of improvements in terms of thermostability. One deletion mutant termed D3 (residues(686-688)) was obtained and exhibited enhanced thermostability due to deletion of the flexible region at the C-terminus. The combination of the two strategies yielded the mutant M18 (Y477A/D3/Y175C/L215P/R473E). It retained 66% of its initial activity after incubation at 60 degrees C for 72 hrs, whereas that of the wild-type enzyme was only 35%. After incubation at 65 degrees C for 4 h, M18 retained 50.6% of its initial activity, whereas that of the wild-type was only 16.8%, respectively. Additionally, kinetic studies revealed that the K-m of M17 (Y477A/D3/Y175C/L215P) was decreased by 33.9% and that the K-cat/K-m value of M17 increased by 50%, while M18 exhibited K-m and K-cat/K-m values that were similar to those of the wild-type enzyme. The attractive improved thermostability and the high catalytic efficiency made M17 and M18 more suitable for industrial application. (C) 2015 Elsevier B. V. All rights reserved. |
| WOS标题词 | Science & Technology ; Life Sciences & Biomedicine |
| 类目[WOS] | Biotechnology & Applied Microbiology |
| 研究领域[WOS] | Biotechnology & Applied Microbiology |
| 关键词[WOS] | SITE-DIRECTED MUTAGENESIS ; BIOCHEMICAL-CHARACTERIZATION ; ANGSTROM RESOLUTION ; THERMOTOGA-MARITIMA ; CRYSTAL-STRUCTURE ; I PULLULANASE ; STABILITY ; CLONING ; PROTEINS ; ENZYMES |
| 收录类别 | SCI |
| 语种 | 英语 |
| WOS记录号 | WOS:000360650000002 |
| 源URL | [http://124.16.173.210/handle/834782/1521]  |
| 专题 | 天津工业生物技术研究所_基因工程与微生物应用技术研究组 宋恢_期刊论文 |
| 作者单位 | Chinese Acad Sci, Ind Enzymes Natl Engn Lab, Tianjin Inst Ind Biotechnol, Tianjin 300308, Peoples R China |
| 推荐引用方式 GB/T 7714 | Li, Shu-fang,Xu, Jian-yong,Bao, Yun-juan,et al. Structure and sequence analysis-based engineering of pullulanase from Anoxybacillus sp LM18-11 for improved thermostability[J]. JOURNAL OF BIOTECHNOLOGY,2015,210:8-14. |
| APA | Li, Shu-fang,Xu, Jian-yong,Bao, Yun-juan,Zheng, Hong-chen,&Song, Hui.(2015).Structure and sequence analysis-based engineering of pullulanase from Anoxybacillus sp LM18-11 for improved thermostability.JOURNAL OF BIOTECHNOLOGY,210,8-14. |
| MLA | Li, Shu-fang,et al."Structure and sequence analysis-based engineering of pullulanase from Anoxybacillus sp LM18-11 for improved thermostability".JOURNAL OF BIOTECHNOLOGY 210(2015):8-14. |
入库方式： OAI收割
来源：天津工业生物技术研究所
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