1.4.5 Enzyme Sources
The large majority of enzymes used for biotransformations in organic chemistry are
employed in a crude form and are relatively inexpensive. Many preparations
typically contain only about 1–30% of actual enzyme, the remainder being inactive
proteins, stabilizers, buffer salts, or carbohydrates from the fermentation broth from
which they have been isolated. This may even be an advantage, because crude
preparations are often more stable than purified enzymes.
Pure enzymes are usually very expensive and are mostly sold by the unit, while
crude preparations are often shipped in kg amounts. Since the techniques for protein
purification through His- or Strep-tagging are becoming easier and more economic,
the use of (partially) purified enzymes in biotransformations is rapidly increasing.
The main sources of enzymes for biotransformations are as follows [149, 150]:
– The detergent industry produces many proteases and lipases in huge amounts.
These are used as additives for detergents to effect the hydrolysis of
proteinogenic and fatty impurities in the laundry process at neutral pH and
modest temperatures.
– The food industry uses proteases and lipases for meat and cheese processing and
for the transesterification of fats and oils [151]. Glycosidases and decarboxylases
are predominantly employed in the brewing and baking industries, respectively.
– Numerous enzymes can be isolated from cheap mammalian organs, such as
kidney or liver, or from slaughter waste.
– Only a small proportion of enzymes used in biotransformations is obtained from
plant sources, such as fruits (e.g., fig, papaya, pineapple) and vegetables (e.g.,
tomato, potato). While sensitive plant cell cultures were used in the past, they are
nowadays cloned into a sturdy host microorganism (Escherichia coli, Pichia,
Aspergillus) for their production.
– The richest and most convenient sources of enzymes are microorganisms. An
impressive number of biocatalysts are derived from bacterial and (lower) fungal
origin by cheap fermentation.
Primary and secondary metabolism The following guidelines help to enhance
the hit-rate in search of a suitable (microbial) enzyme for the transformation of
non-natural compounds: [152]
The central (primary) metabolism provides energy to sustain life and growth by
breakdown of carbon sources, e.g. via glycolysis of a carbohydrate yielding acetylCoA, which is burnt in the Krebs-cycle to CO 2 and H 2 O. In order to optimize the
metabolic flux, enzymes from primary metabolic pathways always have been under
heavy selection pressure to improve their catalytic efficiency and specificity for
their substrates. Although they appear ideal for preparative-scale biotransformations regarding their catalytic rates (~100 s
À1 ), they possess a very narrow substrate
spectrum. In other words, they are fast, but only on a single natural substrate. In
contrast, for the biotransformation of non-natural compounds, catalytic allrounders
are required, which are predominantly found in secondary metabolic pathways
1.4 Enzyme Properties and Nomenclature
25
The large majority of enzymes used for biotransformations in organic chemistry are
employed in a crude form and are relatively inexpensive. Many preparations
typically contain only about 1–30% of actual enzyme, the remainder being inactive
proteins, stabilizers, buffer salts, or carbohydrates from the fermentation broth from
which they have been isolated. This may even be an advantage, because crude
preparations are often more stable than purified enzymes.
Pure enzymes are usually very expensive and are mostly sold by the unit, while
crude preparations are often shipped in kg amounts. Since the techniques for protein
purification through His- or Strep-tagging are becoming easier and more economic,
the use of (partially) purified enzymes in biotransformations is rapidly increasing.
The main sources of enzymes for biotransformations are as follows [149, 150]:
– The detergent industry produces many proteases and lipases in huge amounts.
These are used as additives for detergents to effect the hydrolysis of
proteinogenic and fatty impurities in the laundry process at neutral pH and
modest temperatures.
– The food industry uses proteases and lipases for meat and cheese processing and
for the transesterification of fats and oils [151]. Glycosidases and decarboxylases
are predominantly employed in the brewing and baking industries, respectively.
– Numerous enzymes can be isolated from cheap mammalian organs, such as
kidney or liver, or from slaughter waste.
– Only a small proportion of enzymes used in biotransformations is obtained from
plant sources, such as fruits (e.g., fig, papaya, pineapple) and vegetables (e.g.,
tomato, potato). While sensitive plant cell cultures were used in the past, they are
nowadays cloned into a sturdy host microorganism (Escherichia coli, Pichia,
Aspergillus) for their production.
– The richest and most convenient sources of enzymes are microorganisms. An
impressive number of biocatalysts are derived from bacterial and (lower) fungal
origin by cheap fermentation.
Primary and secondary metabolism The following guidelines help to enhance
the hit-rate in search of a suitable (microbial) enzyme for the transformation of
non-natural compounds: [152]
The central (primary) metabolism provides energy to sustain life and growth by
breakdown of carbon sources, e.g. via glycolysis of a carbohydrate yielding acetylCoA, which is burnt in the Krebs-cycle to CO 2 and H 2 O. In order to optimize the
metabolic flux, enzymes from primary metabolic pathways always have been under
heavy selection pressure to improve their catalytic efficiency and specificity for
their substrates. Although they appear ideal for preparative-scale biotransformations regarding their catalytic rates (~100 s
À1 ), they possess a very narrow substrate
spectrum. In other words, they are fast, but only on a single natural substrate. In
contrast, for the biotransformation of non-natural compounds, catalytic allrounders
are required, which are predominantly found in secondary metabolic pathways
1.4 Enzyme Properties and Nomenclature
25