Food Chemistry

124 2 Enzymes

Fig. 2.26.Plotting slopes (a) and ordinate intercepts (b)
from Fig. 2.25 versus 1/[A 0 ]

Fig. 2.27.Evaluation of a two-substrate reaction, pro-
ceeding through a binary enzyme-substrate complex
(according toLineweaverandBurk).[A 0 ] 4 >[A 0 ] 3 >
[A 0 ] 2 >[A 0 ] 1

of each other in catalysis, theMichaelis–Menten
kinetics, as outlined under sections 2.5.1.1
and 2.5.1.2, are valid. However, when the
subunits cooperate, the enzymes deviate from
these kinetics. This is particularly true in the
case of positive cooperation when the enzyme is

activated by the substrate. In this kind of plot, v 0
versus[A 0 ]yields not a hyperbolic curve but a sat-
uration curve with a sigmoidal shape (Fig. 2.28).

Thus, enzymes which do not obey theMichaelis–

Menten model of kinetics are allosterically

regulated. These enzymes have a site which

reversibly binds the allosteric regulator (sub-

strate, cosubstrate or low molecular weight

compound) in addition to an active site with

a binding and transforming locus. Allosteric

enzymes are, as a rule, engaged at control sites of

metabolism. An example is tetrameric phospho-

fructokinase, the key enzyme in glycolysis. In

glycolysis and alcoholic fermentation it catalyzes

the phosphorylation of fructose-6-phosphate

to fructose-1,6-diphosphate. The enzyme is

activated by its substrate in the presence of

ATP. The prior binding of a substrate molecule

which enhances the binding of each succeeding

substrate molecule is called positive cooperation.

The two enzyme-catalyzed reactions, one which

obeysMichaelis–Mentenkinetics and the other

which is regulated by allosteric effects, can be

reliably distinguished experimentally by com-

paring the ratio of the substrate concentration

needed to obtain the observed value of 0.9V to

that needed to obtain 0.1 V. This ratio, denoted

as Rs, is a measure of the cooperativity of the

interaction.

RS=

(A 0 ) 0 .9V

(A 0 ) 0 .1V

(2.63)

For all enzymes which obeyMichaelis–Menten

kinetics, Rs=81 regardless of the value of Km

or V. The value of Rsis either lower or higher

than 81 for allosteric enzymes. Rs<81 is

Fig. 2.28.The effect of substrate concentration on the

catalytic reaction rate.aEnzyme obeyingMichaelis–

Mentenkinetics;ballosterically regulated enzyme with

positive cooperativity;callosterically regulated en-

zyme with negative cooperativity