|Budget Amount *help
¥7,900,000 (Direct Cost : ¥7,900,000)
Fiscal Year 1996 : ¥800,000 (Direct Cost : ¥800,000)
Fiscal Year 1995 : ¥7,100,000 (Direct Cost : ¥7,100,000)
I.Coenzyme collecting column system
We extablished optimal conditions for collecting coenzymes.
1. Optimal conditons for collecting NAD^+ or NADH
Adsorption : 50mM Tris-HCl, pH8.5, Elution : 50mM MES-NaOH,pH6.0
2. Optimal conditions for collecting ATP or AMP
Adsorption : 50mM GlyGly-NaOH,pH9.5, Elution : 50mM Citrate, pH4.0
3. Optimal conditions for collecting ADP
Adsorption : 50mM CHS-NaOH,pH9.5, Elution : 50mM Citrate, pH4.0
II.Model systems for consumption and regeneration of coenzymes.
We immobilized enzymes for consuming and regenerating NADH or ATP using AffiGE115.
1. Reaction system for NAD^+/NADH
Immobilizing condition for L-lactate dehydrogenase (LDH) : 100mMMOPS-NaOH,pH6.0
Immobilizing condition for formate dehydrogenase (FDH) : 100mM MOPS-NaOH,pH8.0
2. Reaction system for ATP/ADP
Immobilizing condition for hexokinase (HK) : 100mM MOPS-NaOH,pH6.0
Immobilizing condition for pyruvate kinase (PK) : 100mM HEPES,pH8.0
3. Stability of immobilized enzymes
Immobilized FDH had kept 80% of its activity after one month. Activities of other immobilized enzymes somewhat increased during storage.