210
M. Rueping, E. Sugiono
mate with the use of nicotineamide dinucleotide as hydride donor
(Eq. 1).
2-ketogluarate + NH 3 + NAD(P)H glutamate + NAD(P)
+
(1)
The reaction is reversible, although the equilibrium lies in the direction of glutamate formation. Likewise, other amino acid dehydrogenases assimilate ammonia analogous to the GDH reaction, but are
specific for other keto acid/amino acid substrate pairs, e.g. alanine dehydrogenase catalyzes the reductive amination of pyruvate to alanine.
Typically, a high activity of these dehydrogenases is observed if a high
concentration of ammonia is in the environment or under stress conditions such as high temperature, salinity, senescence, or environmental
pollution.
A mechanism for the catalytic activity of dehydrogenases has been
proposed following detailed kinetic studies and crystallographic structures. One important feature in the reductive amination of the 2-ketoglutarate is the protonation of the α-iminoglutarate by aspartate D165,
resulting in the formation of an imium-ion. Subsequent hydride transfer
from the nicotineamide dinucleotide leads to the formation of the amino
acid glutamate (Fig. 1).
To support this mechanism the putative catalytic aspartyl residue
D165 in the active site of GDH has been replaced with other amino
acids, such as alanine, serine or histidine by site directed mutagenesis.
Although the modified enzymes still appeared to be correctly folded the
high catalytic activity was lost, indicating the vital role of the amino
acid D165 for its catalytic activity.
Based on the above activation mechanism we wondered whether it
would be possible to develop a biomimetic, organocatalytic reductive
amination or transfer hydrogenation of ketimines. We reasoned that the
activation of the imine by catalytic protonation through the Brønsted
acid should enable the hydrogen transfer from a suitable NADH mimic
to yield the corresponding amine (Fig. 2). Hence, initial experiments
focused on the examination of various Brønsted acids in combination
with different hydride sources (Rueping et al. 2005a).
From this survey we found that several proton acids catalyze the
reduction of ketimines using Hantzsch dihydropyridine 2 as the hydride donor. With regard to the different acids tested diphenyl phosphate
M. Rueping, E. Sugiono
mate with the use of nicotineamide dinucleotide as hydride donor
(Eq. 1).
2-ketogluarate + NH 3 + NAD(P)H glutamate + NAD(P)
+
(1)
The reaction is reversible, although the equilibrium lies in the direction of glutamate formation. Likewise, other amino acid dehydrogenases assimilate ammonia analogous to the GDH reaction, but are
specific for other keto acid/amino acid substrate pairs, e.g. alanine dehydrogenase catalyzes the reductive amination of pyruvate to alanine.
Typically, a high activity of these dehydrogenases is observed if a high
concentration of ammonia is in the environment or under stress conditions such as high temperature, salinity, senescence, or environmental
pollution.
A mechanism for the catalytic activity of dehydrogenases has been
proposed following detailed kinetic studies and crystallographic structures. One important feature in the reductive amination of the 2-ketoglutarate is the protonation of the α-iminoglutarate by aspartate D165,
resulting in the formation of an imium-ion. Subsequent hydride transfer
from the nicotineamide dinucleotide leads to the formation of the amino
acid glutamate (Fig. 1).
To support this mechanism the putative catalytic aspartyl residue
D165 in the active site of GDH has been replaced with other amino
acids, such as alanine, serine or histidine by site directed mutagenesis.
Although the modified enzymes still appeared to be correctly folded the
high catalytic activity was lost, indicating the vital role of the amino
acid D165 for its catalytic activity.
Based on the above activation mechanism we wondered whether it
would be possible to develop a biomimetic, organocatalytic reductive
amination or transfer hydrogenation of ketimines. We reasoned that the
activation of the imine by catalytic protonation through the Brønsted
acid should enable the hydrogen transfer from a suitable NADH mimic
to yield the corresponding amine (Fig. 2). Hence, initial experiments
focused on the examination of various Brønsted acids in combination
with different hydride sources (Rueping et al. 2005a).
From this survey we found that several proton acids catalyze the
reduction of ketimines using Hantzsch dihydropyridine 2 as the hydride donor. With regard to the different acids tested diphenyl phosphate