In competitive inhibition of
enzyme catalysis, binding of an inhibitor prevents binding of the target molecule of the enzyme, also known as the substrate. This is accomplished by blocking the binding site of the substrate – the active site – by some means. The Vmax indicates the maximum velocity of the reaction, while the Km is the amount of substrate needed to reach half of the Vmax. Km also plays a part in indicating the tendency of the substrate to bind the enzyme. Competitive inhibition can be overcome by adding more substrate to the reaction, which increases the chances of the enzyme and substrate binding. As a result, competitive inhibition alters only the Km, leaving the Vmax the same. Most competitive inhibitors function by binding reversibly to the active site of the enzyme. This, however, is a misleading
oversimplification, as there are many possible mechanisms by which an enzyme may bind either the inhibitor or the substrate but never both at the same time. At any given moment, the enzyme may be bound to the inhibitor, the substrate, or neither, but it cannot bind both at the same time. During competitive inhibition, the inhibitor and substrate compete for the active site. The active site is a region on an enzyme to which a particular protein or substrate can bind. The active site will thus only allow one of the two complexes to bind to the site, either allowing a reaction to occur or yielding it. In competitive inhibition, the inhibitor resembles the substrate, taking its place and binding to the active site of an enzyme. Increasing the substrate concentration would diminish the "competition" for the substrate to properly bind to the active site and allow a reaction to occur. When the substrate is of higher concentration than the concentration of the competitive inhibitor, it is more probable that the substrate will come into contact with the enzyme's active site than with the inhibitor's. Competitive inhibitors are commonly used to make pharmaceuticals. An example of non-drug related competitive inhibition is in the prevention of browning of fruits and vegetables. For example,
tyrosinase, an enzyme within mushrooms, normally binds to the substrate,
monophenols, and forms brown o-quinones. Competitive substrates, such as 4-substituted benzaldehydes for mushrooms, compete with the substrate lowering the amount of the monophenols that bind. These inhibitory compounds added to the produce keep it fresh for longer periods of time by decreasing the binding of the monophenols that cause browning. In competitive inhibition, the maximum velocity (V_\max) of the reaction is unchanged, while the apparent affinity of the substrate to the binding site is decreased (the K_d dissociation constant is apparently increased). The change in K_m (
Michaelis–Menten constant) is parallel to the alteration in K_d, as one increases the other must decrease. When a competitive inhibitor is bound to an enzyme the K_m increases. This means the binding affinity for the enzyme is decreased, but it can be overcome by increasing the concentration of the substrate. Any given competitive inhibitor concentration can be overcome by increasing the substrate concentration. In that case, the substrate will reduce the availability for an inhibitor to bind, and, thus, outcompete the inhibitor in binding to the enzyme. MPTP is biologically activated by MAO-B, an isozyme of
monoamine oxidase (MAO) which is mainly concentrated in neurological disorders and diseases. Later, it was discovered that MPTP causes symptoms similar to that of
Parkinson's disease. Cells in the central nervous system (astrocytes) include MAO-B that oxidizes MPTP to 1-methyl-4-phenylpyridinium (MPP+), which is toxic. This prevents the substrate itself from binding which halts the production of folic acid, an essential nutrient. Bacteria must synthesize folic acid because they do not have a transporter for it. Without folic acid, bacteria cannot grow and divide. Therefore, because of sulfa drugs' competitive inhibition, they are excellent antibacterial agents. An example of competitive inhibition was demonstrated experimentally for the enzyme succinic dehydrogenase, which catalyzes the oxidation of
succinate to
fumarate in the
Krebs cycle.
Malonate is a competitive inhibitor of succinic dehydrogenase. The binding of succinic dehydrogenase to the substrate, succinate, is competitively inhibited. This happens because malonate's chemistry is similar to succinate. Malonate's ability to inhibit binding of the enzyme and substrate is based on the ratio of malonate to succinate. Malonate binds to the active site of succinic dehydrogenase so that succinate cannot. Thus, it inhibits the reaction.
Equation The Michaelis–Menten Model can be an invaluable tool to understanding enzyme kinetics. According to this model, a plot of the reaction velocity (V0) associated with the concentration [S] of the substrate can then be used to determine values such as Vmax, initial velocity, and Km (Vmax/2 or affinity of enzyme to substrate complex). Competitive inhibition increases the apparent value of the
Michaelis–Menten constant, K^\text{app}_m, such that initial rate of reaction, V_0, is given by : V_0 = \frac{V_\max\,[S]}{K^\text{app}_m + [S]} where K^\text{app}_m=K_m(1+[I]/K_i), K_i is the inhibitor's dissociation constant and [I] is the inhibitor concentration. V_\max remains the same because the presence of the inhibitor can be overcome by higher substrate concentrations. K^\text{app}_m, the substrate concentration that is needed to reach V_\max / 2, increases with the presence of a competitive inhibitor. This is because the concentration of substrate needed to reach V_\max with an inhibitor is greater than the concentration of substrate needed to reach V_\max without an inhibitor.
Derivation In the simplest case of a single-substrate enzyme obeying Michaelis–Menten kinetics, the typical scheme : E + S [k_1][k_{-1}] ES ->[k_2] E + P is modified to include binding of the inhibitor to the free enzyme: : EI + S [k_{-3}][k_3] E + S + I [k_1][k_{-1}] ES + I ->[k_2] E + P + I Note that the inhibitor does not bind to the ES complex and the substrate does not bind to the EI complex. It is generally assumed that this behavior is indicative of both compounds binding at the same site, but that is not strictly necessary. As with the derivation of the Michaelis–Menten equation, assume that the system is at steady-state, i.e. the concentration of each of the enzyme species is not changing. : \frac{d[\ce E]}{dt} = \frac{d[\ce{ES}]}{dt} = \frac{d[\ce{EI}]}{dt} = 0. Furthermore, the known total enzyme concentration is [\ce E]_0 = [\ce E] + [\ce{ES}] + [\ce{EI}], and the velocity is measured under conditions in which the substrate and inhibitor concentrations do not change substantially and an insignificant amount of product has accumulated. We can therefore set up a system of equations: {{NumBlk|:| [\ce E]_0 = [\ce E] + [\ce{ES}] + [\ce{EI}]|}} {{NumBlk|:| \frac{d[\ce E]}{dt} = 0 = -k_1[\ce E][\ce S] + k_{-1}[\ce{ES}] + k_2[\ce{ES}] -k_3[\ce E][\ce I] + k_{-3}[\ce{EI}] |}} {{NumBlk|:| \frac{d[\ce{ES}]}{dt} = 0 = k_1[\ce E][\ce S] - k_{-1}[\ce{ES}] - k_2[\ce{ES}] |}} {{NumBlk|:| \frac{d[\ce{EI}]}{dt} = 0 = k_3[\ce E][\ce I] - k_{-3}[EI] |}} where [S], [I] and [E]_0 are known. The initial velocity is defined as V_0 = d[\ce P]/dt = k_2 [\ce{ES}], so we need to define the unknown [ES] in terms of the knowns [S], [I] and [E]_0. From equation (), we can define
E in terms of
ES by rearranging to : k_1[\ce E][\ce S]=(k_{-1}+k_2)[\ce{ES}] Dividing by k_1[\ce S] gives : [\ce E] = \frac{(k_{-1}+k_2)[\ce{ES}]}{k_1[\ce S]} As in the derivation of the Michaelis–Menten equation, the term (k_{-1}+k_2)/k_1 can be replaced by the macroscopic rate constant K_m: {{NumBlk|:| [\ce E] = \frac{K_m[\ce{ES}]}\ce{[S]} |}} Substituting equation () into equation (), we have : 0 = \frac{k_3[\ce I]K_m[\ce{ES}]}\ce{[S]} - k_{-3}[\ce{EI}] Rearranging, we find that : [\ce{EI}] = \frac{K_m k_3[\ce I][\ce{ES}]}{k_{-3}[\ce S]} At this point, we can define the dissociation constant for the inhibitor as K_i = k_{-3}/k_3, giving {{NumBlk|:| [\ce{EI}] = \frac{K_m[\ce I][\ce{ES}]}{K_i[\ce S]}|}} At this point, substitute equation () and equation () into equation (): : [\ce E]_0 = \frac{K_m[\ce{ES}]}\ce{[S]} + [\ce{ES}] + \frac{K_m[\ce I][\ce{ES}]}{K_i[\ce S]} Rearranging to solve for ES, we find : [\ce E]_0 = [\ce{ES}] \left ( \frac{K_m}\ce{[S]} + 1 + \frac{K_m[\ce I]}{K_i[\ce S]} \right )= [\ce{ES}] \frac{K_m K_i + K_i[\ce S] + K_m[\ce I]}{K_i[\ce S]} {{NumBlk|:| [\ce{ES}] = \frac{K_i [\ce S][\ce E]_0}{K_m K_i + K_i[\ce S] + K_m[\ce I]}|}} Returning to our expression for V_0, we now have: : V_0 = k_2[\ce{ES}] = \frac{k_2 K_i [\ce S][\ce E]_0}{K_m K_i + K_i[\ce S] + K_m[\ce I]} : V_0 = \frac{k_2 [\ce E]_0 [\ce S]}{K_m + [\ce S] + K_m\frac{[\ce I]}{K_i}} Since the velocity is maximal when all the enzyme is bound as the enzyme-substrate complex, V_\max = k_2 [\ce E]_0. Replacing and combining terms finally yields the conventional form: {{NumBlk|:| V_0 = \frac{V_{\max}[\ce S]}{K_m\left(1 + \frac{[\ce I]}{K_i}\right) + [\ce S]}|}} To compute the concentration of competitive inhibitor [I] that yields a fraction f_{V{_0}} of velocity V_0 where 0 : {{NumBlk|:| [\ce I]= \left(\frac{1}{f_{V{_0}}} -1\right)K_i\left(1+\frac{[\ce S]}{K_m}\right)|}} == See also ==