Introduction to Enzyme and Coenzyme Chemistry - E-Library Home

Enzymatic Hydrolysis and Group Transfer Reactions 95
Zn 2+
H 3 C
HO
HO
O
O
OH
O − P NH
O
H
O O
Glu 143
O
NH-Trp
Figure 5.15 Inhibition of thermolysin by
phosphoramidon.
active site zinc, acting as an analogue of the oxyanion tetrahedral intermediate,
and is therefore bound extremely tightly by the enzyme. Further phosphonamidate
inhibitors have now been devised and synthesised which bind even
more tightly.
A further class of metalloproteases contain two divalent metal ions,
arranged in a binuclear metal cluster. One example of this family of enzymes
is leucine aminopeptidase. In these enzymes it is thought that one metal centre
activates the nucleophilic water molecule, while the second metal centre activates
the amide carbonyl group via Lewis acid activation.
The aspartyl proteases
The aspartyl (or acid) proteases are characterised by the presence of two active
site aspartate residues whose carboxylate side chains are involved in catalysis.
They are probably the smallest class of protease enzymes, but have recently
come to prominence through the discovery that the HIV-1 virus contains an
essential aspartyl protease enzyme, which we shall examine below. The aspartyl
proteases are characteristic in their ability to operate at low pH: the enzyme
pepsin operates in the range 2–4, which makes it well suited for operating in the
acidic environment of the stomach. This pH requirement will become apparent
when we examine the mechanism of these enzymes, since one of the two
aspartate residues must be protonated for activity.
This family of enzymes are also characterised by their inhibition by
low levels (1 mgml ) of pepstatin (Figure 5.16), a modiWcation of a naturally
occurring peptide containing the unusual amino acid statine (see Figure 5.20 for
an illustration of the mechanism of inhibition by this class of inhibitor).
Val-Val-Val
H
N
OH
O
NH-Ala-Statine
O
statine
Figure 5.16 Pepstatin.