Enzyme Technology
Use of 'unnatural' substrates
Many enzymes are not totally specific for their natural substrates. Some have
been found to catalyse reactions quite different from those given as the normal
reactions and reflected in their name and EC number. Sometimes it necessary to
place the enzyme in an unusual environment in order to display new activities.
Thus, lipases act as transesterases in primarily non-aqueous environments (see
Chapter 7). In other cases, changes in the environment are not necessary.
Glucose oxidase (see reaction scheme [1.1]) is specific for its reducing
substrate (D-glucose) but fairly non-specific in its choice of oxidant, normally
molecular oxygen. It has been established that benzoquinone is also an effective
electron-accepting substrate:
[8.1]
b-D-glucose +
benzoquinone D-glucono-1,5-lactone + hydroquinone
The product of this reaction, hydroquinone, is a valuable organic chemical,
being used in the photographic industry and as an antioxidant. The reaction
gives nearly 100% yields, with no possibility of the peroxide-induced
inactivation which occurs using molecular oxygen as oxidant. Because of the
ready solubility of benzoquinone and low solubility of molecular oxygen, the
above reaction ([8.1]) can give productivity rates several times greater than
the 'natural' reaction ([1.1]).
Acetylcholinesterase (EC 3.1.1.7) normally catalyzes the hydrolysis of
acetylcholine, the excitatory neurotransmitter, in the synaptic junctions of
vertebrates.
[8.2]
acetylcholine + water
choline + acetic acid
The acetylcholinesterase from the electric eel has been found additionally to
catalyse the stereospecific hydrolysis of acetyl-D-carnitine but not
acetyl-L-carnitine.
[8.3]
acetyl-D-carnitine + water
D-Carnitine + acetic acid
Although the reaction involving acetyl-D-carnitine has a second-order rate
(specificity) constant four orders of magnitude smaller than that utilising
acetylcholine, the productivities at high substrate concentrations are
comparable, the 'natural' substrate, acetylcholine, causing pronounced substrate
inhibition which is not apparent with acetyl-D-carnitine. The 'unnatural'
reaction is useful as it may be used in the preparation of L-carnitine, one of
the vitamins that has numerous therapeutic applications; the D-isomer being
biologically inactive. The enzyme can, therefore, be used in a manner similar to
the way in which aminoacylase is used to resolve racemic amino acids. Chemically
synthesised racemic DL-carnitine may be acetylated, using acetyl chloride, to
give acetyl-DL-carnitine. The acetylcholinesterase may then be used to produce a
mixture of acetyl-L-carnitine and D-carnitine that may be simply resolved by
ion-exchange chromatography. The acetyl-L-carnitine is as biologically active as
L-carnitine and may be used directly.
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This page was established in 2004 and last updated by Martin
Chaplin on
6 August, 2014
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