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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. 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:
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b-D-glucose + benzoquinone
 D-glucono-1,5-lactone + hydroquinone
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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 catalyses the
hydrolysis of acetylcholine, the excitatory neurotransmitter, in
the synaptic junctions of vertebrates. |
 The acetylcholinesterase from the electric eel has been found additionally to
catalyse the stereospecific hydrolysis of acetyl-D-carnitine but not
acetyl-L-carnitine.
| 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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