2009_Book_FoodChemistry

2.4 Theory of Enzyme Catalysis 115
only react with substrate by donating an electron
pair (nucleophilic catalysis). Electrophilic reactions
occur mostly by involvement of carbonyl
groups (example 7, Table 2.8) or with the help of
metal ions.
A number of peptidase and esterase enzymes react
covalently in substitution reactions by a twostep
nucleophilic mechanism. In the first step, the
enzyme is acylated; in the second step, it is deacylated.
Chymotrypsin will be discussed as an example
of this reaction mechanism. Its activity is
dependent on His 57 and Ser 195 , which are positioned
in close proximity within the active site
of the enzyme because of folding of the peptide
chain (Fig. 2.16).
Because Asp 102 is located in hydrophobic surroundings,
it can polarize the functional groups in
close proximity to it. Thus, His 57 acts as a strong
general base and abstracts a proton from the
OH-group of the neighboring Ser 195 residue (step
‘a’, Fig. 2.17). The oxygen remaining on Ser 195
thus becomes a strong nucleophile and attacks
the carbon of the carbonyl group of the peptide
bond of the substrate. At this stage an amine (the
first product) is released (step ‘b’, Fig. 2.17) and
the transient covalently-bound acyl enzyme is
formed. A deacylation step follows. The previous
position of the amine is occupied by a water
molecule. Again, His 57 , through support from
Asp 102 , serves as a general base, abstracting the
proton from water (step ‘c’, Fig. 2.17). This is
followed by nucleophilic attack of the resultant
OH ⊖ ion on the carbon of the carbonyl group of
the acyl enzyme (step ‘d’, Fig. 2.17), resulting
Fig. 2.16. Polypeptide chain conformation in the
chymotrypsin molecule (according to Lehninger, 1977)
Fig. 2.17. Postulated reaction mechanism for chymotrypsin
activity (according to Blow et al., 1969)
in free enzyme and the second product of the
enzymic conversion.
An exceptionally reactive serine residue has
been identified in a great number of hydrolase
enzymes, e. g., trypsin, subtilisin, elastase, acetylcholine
esterase and some lipases. These
enzymes appear to hydrolyze their substrates
by a mechanism analogous to that of chymotrypsin.
Hydrolases such as papain, ficin and
bromelain, which are distributed in plants, have
a cysteine residue instead of an “active” serine
residue in their active sites. Thus, the transient
intermediates are thioesters.
Enzymes involved in the cleavage of carbohydrates
can also function by the above mechanism.
Figure 2.18 shows that amylose hydrolysis by
β-amylase occurs with the help of four functional
groups in the active site. The enzyme-substrate
complex is subjected to a nucleophilic attack