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Conversion of ammonia or urea into essential amino acids, L‐leucine, L‐valine, and L‐isoleucine, using artificial cells containing an immobilized multienzyme system and dextran‐NAD+. 2. Yeast alcohol dehydrogenase for coenzyme recycling
Author(s) -
Gu KF,
Chang TM
Publication year - 1990
Publication title -
biotechnology and applied biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.468
H-Index - 70
eISSN - 1470-8744
pISSN - 0885-4513
DOI - 10.1111/j.1470-8744.1990.tb00094.x
Subject(s) - isoleucine , valine , leucine , urea , chemistry , chromatography , biochemistry , alcohol dehydrogenase , substrate (aquarium) , amino acid , dehydrogenase , alcohol , biology , enzyme , ecology
Semipermeable nylon‐polyethylenimine artificial cells containing leucine dehydrogenase (EC 1.4.1.9), alcohol dehydrogenase (EC 1.1.1.1), urease (EC 3.5.1.5), and dextran‐NAD+ were prepared. Artificial cells could convert ammonia or urea into L‐leucine, L‐valine, and L‐isoleucine. For batch conversion in 20.0 mM of ammonium acetate substrate solutions, in 2 h 0.2 ml of artificial cells could produce 4.48 mumol of L‐leucine, 9.98 mumol of L‐valine, or 5.96 mumol of L‐isoleucine. The corresponding conversion ratios were 22.4, 49.9, and 29.8%. In 20.0 mM of urea substrate solutions, 13.71 mumol of L‐leucine, 16.12 mumol of L‐valine, or 13.44 mumol of L‐isoleucine was produced and the conversion ratios were 68.6, 80.6, and 67.2%. The substrate specificity of leucine dehydrogenase for the reductive amination was determined. Of the three branched‐chain amino acids produced, the production rates of L‐valine were the highest. The apparent Km values were as follows: 0.32 mM for alpha‐ketoisocaproate, 1.63 mM for alpha‐ketoisovalerate, and 0.73 mM for Dl‐alpha‐keto‐beta‐methyl‐n‐valerate. The leucine dehydrogenase multienzyme system had a good storage stability. It retained 72.0% of the original activity with artificial cells were stored at 4 degrees C for 6 weeks. The optimum conversion pH and temperature were 8.5–9.0 and 35–40 degrees C. The effects of urea and ammonium salts on conversion rate were also studied. The relative activities in ammonium salts solutions were 45.1–75.9% of those in urea solutions.