Cold adaptation of thermostable enzymes by surface loops' mutagenesis
| Project/Area Number |
23658095
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| Research Category |
Grant-in-Aid for Challenging Exploratory Research
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| Allocation Type | Multi-year Fund |
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| Research Field |
Applied biochemistry
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| Research Institution | Fukui Prefectural University |
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Principal Investigator |
HIBI Takao 福井県立大学, 生物資源学部, 教授 (00285181)
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| Co-Investigator(Kenkyū-buntansha) |
ITOH Takafumi 福井県立大学, 生物資源学部, 講師 (10402827)
NISHIYA Yoshiaki 摂南大学, 理工学部, 教授 (70612307)
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| Project Period (FY) |
2011 – 2013
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| Project Status |
Completed (Fiscal Year 2013)
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| Budget Amount *help |
¥3,640,000 (Direct Cost: ¥2,800,000、Indirect Cost: ¥840,000)
Fiscal Year 2013: ¥650,000 (Direct Cost: ¥500,000、Indirect Cost: ¥150,000)
Fiscal Year 2012: ¥910,000 (Direct Cost: ¥700,000、Indirect Cost: ¥210,000)
Fiscal Year 2011: ¥2,080,000 (Direct Cost: ¥1,600,000、Indirect Cost: ¥480,000)
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| Keywords | タンパク質工学 / 酵素 / 分子設計 / 好冷性酵素 / psychrophilic / enzyme / protein engineering / 省エネルギー / 食品 / 洗剤 / 枯草菌 / X線結晶構造解析 / 酵素分析 / バイオセンサ |
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| Research Abstract |
The high activity of psychrophilic enzymes offers potential benefits through substantial energy savings in large-scale chemical processes. Trp279 and Pro287 of Bacillus sp. TB-90 uricase were predicted to be located at hinges of the interface loop II. The optimum temperature of the W279L+P287G mutant decreased from 50 degree to 30 degree without loss of its thermal stability. Here, we determined its crystal structure at a 2.1 angstrom resolution. The residues 284-289 were missing and the other residues in the loop showed higher temperature factors. The flexibility of the loop may be responsible for the successful cold adaptation. The saturation mutagenesis studies on seven residues in the surface loops of alpha-amylase from Bacillus subtilis were also performed. Psychorphilic mutants of five residues were obtained by the screening using the starch degradation assay. They showed 120-300% increases of the activities with having kept thermal stability.
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Report
(4 results)
Research Products
(14 results)
