Conversion of bacterial allosteric L-lactate dehydrogenases to constitutively active enzymes
| Project/Area Number |
23580120
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Multi-year Fund |
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| Section | 一般 |
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| Research Field |
Applied microbiology
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| Research Institution | Tokyo University of Science |
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Principal Investigator |
TAGUCHI Hayao 東京理科大学, 理工学部, 教授 (90188136)
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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 |
¥5,200,000 (Direct Cost: ¥4,000,000、Indirect Cost: ¥1,200,000)
Fiscal Year 2013: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000)
Fiscal Year 2012: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
Fiscal Year 2011: ¥2,340,000 (Direct Cost: ¥1,800,000、Indirect Cost: ¥540,000)
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| Keywords | アロステリック特性 / 乳酸脱水素酵素 |
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| Research Abstract |
For LCLDH, S67E, N68D, E178K and A235K replacements additively increased the FBP-independent activity, reducing the substrate Km values. The X-ray crystallography indicated that these mutations introduced the inter-subunit salt bridges between the Q-axis related subunits. For LPLDH, D68N and D68H replacements consistently changed the hyperbolic substrate saturation curves to the sigmoidal ones. In the case of TCLDH, L67E, N68D, E178K and A235K (or A235R) replacements (Q mutations) only slightly improved substrate Km in the absence of FBP, though they improved Vm values, instead R173Q and R216L replacements (P mutations) markedly improved the substrate Km. The P-axis and Q-axis mutations additively improved the FBP-independent activity and thermal stability. Structural comparison indicated that LCLDH and TCLDH exhibit greatly different structural changes, and therefore require the different designs that introduce the constitutively high catalytic activities.
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Report
(4 results)
Research Products
(30 results)
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[Journal Article] The crystal structure of D-mandelate dehydrogenase reveals its distinct substrate and coenzyme recognition mechanisms from those of 2-ketopantoate reductase2013
Author(s)
Miyanaga, A., Fujisawa, S., Furukawa, N., Arai, K., Nakajima, M., Taguchi, H..
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Journal Title
Biochem. Biophys. Res. Commun.
Volume: 439
Issue: 1
Pages: 109-114
DOI
Related Report
Peer Reviewed
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