Elucidation of Mechanisms and Control of Functions of Versatile Biocatalysts
| Project/Area Number |
17550155
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Chemistry related to living body
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| Research Institution | Okayama University |
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Principal Investigator |
EMA Tadashi Okayama University, Graduate School of Natural Science and Technology, Associate Professor, 大学院自然科学研究科, 助教授 (20263626)
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| Project Period (FY) |
2005 – 2006
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| Project Status |
Completed (Fiscal Year 2006)
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| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2006: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 2005: ¥2,600,000 (Direct Cost: ¥2,600,000)
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| Keywords | enzyme / biocatalyst / enantioselectivity / control / mutation / alcohol / evolution / intermediate for drug |
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| Research Abstract |
1. Rational control of enantioselectivity of lipase
We selected I1e287 as a mutation site to control the enantioselectivity of a Burkholderia cepacia lipase on the basis of a mechanism and X-ray crystal structures. A mutation that increases the steric bulkiness as compared with the wild-type enzyme will suppress the reaction of the slower-reacting enantiomer and improve the enantioselectivity. On the other hand, a mutation that decreases the steric hindrance will facilitate the reaction of the slower-reacting enantiomer and diminish the enantioselectivity. We have succeeded in controlling (both increasing and decreasing) the enantioselectivity of the lipase rationally by mutating only one amino acid residue on the basis of the mechanism.
2. Synthesis of an optically active porphyrin dimer via a lipase-catalyzed reaction and its function
An enantiomerically pure porphyrin alcohol prepared by the lipase-catalyzed kinetic resolution was linked to isophthalic acid to give a porphyrin dimer. The zinc complex of the dimer showed chiral discrimination in NMR.
3. Asymmetric reduction of ketones using recombinant E. coli
We constructed a recombinant E. coli overproducing a carbonyl reductase that shows high enantioselectivity toward a wide range of ketones, and conducted the asymmetric reduction of more than 20 ketones, and 12 alcohols had enantiomeric purities of >98% ee. The reaction conditions for 2,4-octanedione were optimized, and (S)-2-hydroxy-4-octanone was obtained with the productivity of 41 g/L. An optically active intermediate for an important chiral drug was also prepared by this biotransformation.
4. Preparation of the carbonyl reductase with a His-tag
Carbonyl reductases with a His-tag at the N-or C-terminal were constructed and produced by E. coli.
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Report
(3 results)
Research Products
(11 results)
