those based on NMR relaxation dispersion experiments, have provided clear evidence
that a key to the rate of catalysis is the interconversion of a population of protein
conformations (Section 15.2.4). For example, studies with the enzyme dihydrofolate
reductase have identified five conformations involved in the catalytic process that
interconvert at rates that are identical to those previously determined by conventional
kinetic experiments. Similarly, studies with the enzyme triosephosphate isomerase
have shown that the opening and closing of a loop in the catalytic site has a vital
role in the catalytic cycle. The loop has been shown to open and close at a rate of
10 000 s^1 , a value that is the same as the catalytic rate constant. This correlation
between rate-limiting steps in catalysis and rates of conformational change suggests
that as the substrates bind and are converted to products, the population of conform-
ations adjust and act to drive catalysis along the product pathway.Multienzyme complexes
Studies on multienzyme complexes, including tryptophan synthase and carbamyl phos-
phate synthase, have demonstrated that the active site of one enzyme is coupled to that of
the next enzyme in the metabolic sequence by means of allosteric conformational changes.
The reaction products are channelled from one active site to the next by means of an
intermolecular tunnel. In the case of tryptophan synthase, which is a (ab) 2 complex in
which theaandbsubunits catalyse separate reactions, the tunnel is approximately 25 A ̊in
length whereas that in carbamoyl phosphate synthase is approximately 100 A ̊long. The
tunnels protect reactive intermediates from coming in contact with the external environ-
ment and reduce their transit time to the next active site. In the case of both enzymes the
tunnels are formed prior to the binding of the initial substrates but with some other
multienzymecomplexesthetunnelsareformedafterthesubstratesbindtotheactivesite.
Closely related to multienzyme complexes are the megasynthases responsible for the
synthesis of antibiotics such as penicillin and vancomycin. They are large multifunc-
tional enzymes consisting of clusters of active sites known asmodules. They promote a
series of reactions starting with simple organic intermediates that are progressively
converted to the antibiotic. During the synthetic process, the intermediates are tethered
to carrier proteins that shuttle them in sequence to the designated active site (referred
to asclient enzymes) in the module. NMR evidence indicates that the enzymes and
the carrier proteins exist in an ensemble of dynamic conformations and that successful
docking between the two to promote the next synthetic stage relies on the selection of
the correct conformation in which the enzyme’s carrier protein binding site is exposed.15.5 Control of enzyme activity
15.5.1 Control of the activity of individual enzymes
The activity of an enzyme can be regulated in two basic ways:- by alteration of the kinetic conditions under which the enzyme is operating;
- by alteration of the amount of the active form of the enzyme present by promoting
enzyme synthesis, enzyme degradation or the chemical modification of the enzyme.
615 15.5 Control of enzyme activity