Development of single molecule display platform for directed evolution of enzymes
| Project/Area Number |
18K14387
|
|---|
| Research Category |
Grant-in-Aid for Early-Career Scientists
|
| Allocation Type | Multi-year Fund |
|---|
| Review Section |
Basic Section 38030:Applied biochemistry-related
|
|---|
| Research Institution | Nagoya University |
|---|
Principal Investigator |
|
|---|
| Project Period (FY) |
2018-04-01 – 2022-03-31
|
| Project Status |
Completed (Fiscal Year 2021)
|
| Budget Amount *help |
¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000)
Fiscal Year 2020: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000)
Fiscal Year 2019: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000)
Fiscal Year 2018: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
|
|---|
| Keywords | biomolecular engineering / in vitro display / cDNA display / transglutaminase / oxidase / NGS / bioinformatics / substrate profiling / protein engineering / enzyme engineering / D-amino acid oxidase / library screening / directed evolution / in vitro translation / mRNA/cDNA display / in vitro transcription / peroxidases / enzymes |
|---|
| Outline of Final Research Achievements |
We developed a platform for rapid activity-based screening of combinatorial biomolecular libraries during protein engineering, aiming to enable rapid development of modified enzymes and enzymatic substrates for use in industry, diagnostics, and research. Accuracy and in-depth analysis of the selected sequences was enabled by next-generation sequencing and bioinformatics. Unlike conventional methods for protein engineering, this platform takes short time and less labor under acceptable expenses to yield a modified biomolecule with desired properties. Screening and selection system was first optimized using simple model libraries followed by screening of the combinatorial libraries of interest. We have selected very reactive peptide substrates of transglutaminases (TG), TG1 and TG2, and optimized the screening system for the selection of enzymes, e.g. oxidase. Oxidase and transglutaminase are important for industry and diagnostics and for this reason we focused on these two enzymes.
|
| Academic Significance and Societal Importance of the Research Achievements |
選択された生体分子配列の詳細な解析は、酵素の基質特異性や構造-機能相関に関する包括的な知識を提供し、酵素の機能理解や将来の応用設計に重要であることから、科学に貢献する。酵素とその基質の開発は、付加価値の高い製品の効率的かつ持続的な生産、高感度かつ迅速な診断ツール、効率的な治療薬を可能にするため、社会にとって有益である。
|
Report
(5 results)
Research Products
(9 results)
