Introduction to Enzyme and Coenzyme Chemistry - E-Library Home

40 Chapter 3
glycoside hydrolysis (see Section 5.7) also employ an acidic catalytic group to
protonate the glycosidic leaving group – but the enzymatic reaction is some
30 000-fold faster even than the intramolecular reaction, suggesting that the
enzyme is able to further stabilise the transition state for this reaction.
Enzymes also have the ability to carry out bifunctional catalysis: protonation
of the substrate at the same time as deprotonation in another part of
the molecule. An example of bifunctional catalysis is the enzyme ketosteroid
isomerase, whose active site (see Figure 3.14) contains two catalytic residues:
Asp-38 which acts as a catalytic base; and Tyr-14 which acts as an acidic group.
The mechanism (see Figure 3.15) involves the formation of a dienol intermediate
via a concerted step involving simultaneous deprotonation of the substrate
by Asp-14 and protonation of the substrate carbonyl by Tyr-14. In the second
step the tyrosinate group acts as a base, and the substrate is re-protonated by
the protonated Asp-38.
Bifunctional catalysis is thought to make possible the deprotonation of
substrates with apparently high pK a values. Thus, in the above example deprotonation
adjacent to a ketone in solution to form an enolate species would
involve removal of a proton of pK a 18–20, which would be impractical at pH 7.
However, simultaneous protonation to form an enol intermediate makes the
reaction thermodynamically much more favourable.
Figure 3.14 Active site of ketosteroid isomerase (PDB Wle 1E3V). Tyr-14 and Asp-38 are shown
in red.