Industrial Biotransformations

Lactate dehydrogenase
Staphylococcus epidermidis
1) Reaction conditions
[1]: 0.2 M, 35.6 g · L –1 [178.18 g · mol –1 ]
pH: 8.0
T: 30 °C
medium: aqueous
reaction type: redox reaction
catalyst: solubilized enzyme
enzyme: (R)-lactate-NAD oxidoreductase
strain: Staphylococcus epidermidis
CAS (enzyme): [9028–36–8]
2) Remarks
1
COOH
1 = 2-oxo-4-phenyl-butyric acid (OPBA)
2 = 2-hydroxy-4-phenyl-butyric acid (2-HPBA)
E1 = D-lactate dehydrogenase
E2 = formate dehydrogenase
Fig. 1.1.1.28 – 1
● Cofactor regeneration is carried out by formate dehydrogenase from Candida boidinii utilizing
formate and producing CO 2. During cofactor regeneration no by-product is formed that needs
to be separated. In the steady state of the continuously operated stirred tank reactor a total
turnover number of 900 is achieved.
● NAD + is added as cofactor at a concentration of 0.2 mM and ammonium formate at a concentration
of 0.35 M.
● The optimal working pH would have been 6.5, but a pH of 8.0 is chosen due to better solubility
of the hydroxy acid.
● The production is carried out in a continuously operated stirred tank reactor equipped with an
ultrafiltration membrane (cut off 10,000 Da) to retain the enzymes.
● For stabilization of the enzymes 0.15 % mercaptoethanol and 1 mM EDTA are added to the
substrate solution.
● At a conversion of 90 % the pH is shifted from 6.2 (substrate solution) to 8.0. The reason is the
different pK a-values of 2-oxo-4-phenylbutyric acid (pK a = 2.3) and (R)-2-hydroxy-4-phenylbutyric
acid (pK a = 3.76).
● The data given relate to the above process with isolated enzymes.
176
O
CO 2
E1
NADH + H + NAD +
E2
HCOOH
OH
(R)-2
COOH
EC 1.1.1.28