| 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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| 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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