Introduction to Metabolism › Enzymes

Enzyme Inhibition

Notes

Enzyme Inhibition

Sections


Overview

KEY VALUES

  • Vmax
    – Maximal rate of a reaction (every active site bound by substrate)
  • Km
    – Inversely proportional to binding affinity of enzyme to substrate

ENZYME INHIBITION

  • Occurs when a substance reduces activity of an enzyme

Types of inhibition

  • Competitive
    – Substrate & inhibitor compete for active site
    – Greater [S] overcomes inhibition
    – Increases the apparent Km but does NOT affect Vmax
  • Noncompetitive
    – Inhibitor reversibly binds to enzyme outside of active site to deactivate it.
    – Enzymes regain function when inhibitor removed from system
    – Does NOT change Km but lowers Vmax

NOT uncompetitive inhibition in which inhibitors bind enzyme-substrate complexes

  • Uncompetitive inhibition: requires preassembled enzyme-substrate complexes-->more effective when [S] is high
  • Irreversible
    – Inhibitor binds to and permanently deactivates enzyme
    – Only overcome by synthesis of new enzymes

CLINICAL CORRELATION

  • Heavy metals (mercury & lead)
    – Irreversible inhibitors: bind tightly to sulfur groups in enzymes
    – Permanently deactivate them
  • Ethylene glycol (antifreeze) metabolites
    – Toxic to human body
    – Ethanol (competitive inhibitor): used to inhibit alcohol dehydrogenase active site to prevent metabolism of ethylene glycol
  • Angiotension-converting enzyme (ACE) inhibitors
    – Blood pressure lowering agents: noncompetitively inhibit ACE
    – Prevent formation of angiotensin (acts on kidneys to inc. blood pressure)

Full-Length Text

  • Here we will learn enzyme inhibition, which occurs when a substance reduces the activity of an enzyme.
    • The human body uses a complex system of enzyme inhibitors to regulate many processes which are essential for life. - We will use Lineweaver-Burk graphs and their variables to understand the types of inhibition.

First, we will cover the types and definitions of enzyme inhibition.

  • Make a table and list the three main types of inhibition:
    • Competitive
    • Noncompetitive
    • Irreversible
  • Denote that competitive inhibition occurs when the substrate and inhibitor compete for the active site of an enzyme. - Higher substrate concentrations overcome these types of enzymes.
  • Next, denote that noncompetitive inhibition occurs when an inhibitor reversibly binds to an enzyme outside of the active site to deactivate the active site.
    • Noncompetitively inhibited enzymes regain function when the inhibitor is removed from the system.
    • NOTE: Noncompetitive inhibition is not to be confused with uncompetitive inhibition (also called anti-competitive inhibition), which bind to enzyme-substrate complexes to inhibit enzymes.
    • Uncompetitive inhibitors require the enzyme-substrate complexes to be already assembled, and are more effective when substrate concentration is high.
  • Finally, denote that irreversible inhibition occurs when an inhibitor binds to and permanently deactivates an enzyme. - These inhibitors are only overcome by the synthesis of new enzymes.
    • As a clinical correlation, denote that heavy metals, such as mercury and lead, are irreversible inhibitors that bind tightly to sulfur groups on enzymes and permanently deactivate them.

Now, let's address some key definitions.

First, we'll best understand enzyme inhibition when we study its effects on Vmax: the maximal rate of a reaction (which occurs when every enzyme's active site is bound by a substrate) and also Km, which is inversely proportional to the binding affinity of the enzyme to the substrate. As discussed elsewhere, in Lineweaver-Burk plots, use inverse values, so that the graphs are straight lines.

  • Now, also write that:
    • Competitive inhibitors decrease the apparent binding affinity but do NOT affect maximal rate of a reaction. Since Km is inversely proportional to binding affinity, they increase the apparent Km.
    • Noncompetitive inhibitors do NOT affect binding affinity but do decrease the maximal rate of a reaction. (ie, they do NOT change Km, but do lower Vmax).

Let's draw two Lineweaver-Burk graphs to visualize how inhibition changes Vmax and Km in both competitive and noncompetitive inhibition.

  • First, let's draw competitive inhibition.
  • Draw an x-axis.
    • Show the positive and negative coordinates.
  • Now, show the y-axis midway between these values.
  • Label the x-axis, "1/substrate concentration."
  • Label the y-axis, "1/velocity."
  • Draw a diagonal line on the graph to represent a normal enzyme catalytic reaction, with one substrate and enzyme, and without an inhibitor.
    • This line should have a positive slope, as the velocity of the reaction increases with increasing substrate concentration.
  • Label this line, "normal."
  • Label 1/Km at the x-intercept: a high Km means that the enzyme binds the substrate with a low affinity.
  • Label the 1/Vmax at the y-intercept.
  • Now, draw a steeper diagonal line (with the same y-intercept) on the graph to show the same reaction in the presence of a competitive inhibitor.
  • Label this line, "Competitive Inhibitor"
  • Label 1/Km(competitor).
    • Here, the substrate has to compete with the competitive inhibitor for the active site on the enzyme.
    • This creates an apparent increase in Km because of the decreased binding affinity for the substrate. More substrate is now required to produce the same rate of reaction. In other words: when a competitive inhibitor is introduced, the enzyme's apparent affinity for the substrate naturally goes down because there are fewer enzyme molecules available to bind to the substrate (producing an apparent reduction in affinity). In order to reach the same Vmax, more substrate must be added to overwhelm the inhibitor and get to the enzyme active site. Because Km is inversely proportional to binding affinity, when the apparent binding affinity goes DOWN, the apparent Km INCREASES. Thus, on the graph, -1/Km gets closer to zero.
  • Notice that 1/Vmax does not change in the presence of a competitive inhibitor, because the competitive inhibitor does not change the maximal rate at which the reaction can occur (Vmax).
    • High substrate concentrations can overcome this kind of enzyme inhibitor so that all the active sites of the enzyme are bound by substrate.
  • As a clinical correlation, write that the breakdown products of ethylene glycol (antifreeze) are toxic to the human body.
    • Thus, we use the competitive inhibitor ethanol to inhibit the active site on alcohol dehydrogenase.
    • This prevents metabolism of ethylene glycol (prevents generation of its toxic breakdown products).

Now we will draw a second Lineweaver-Burk graph to understand noncompetitive inhibition.

  • Draw an x-axis.
    • Show the positive and negative coordinates.
  • Now, show the y-axis in relation to these values.
  • Label the y-axis: 1/velocity.
  • Label the x-axis: 1/substrate concentration.
  • Draw a diagonal line on the graph to represent a normal enzyme catalytic reaction with one substrate and an enzyme without an inhibitor.
  • Label 1/Km at the x-intercept.
  • Label 1/Vmax at the y-intercept.

Now draw the reaction plot showing the effects of a noncompetitive inhibitor.

  • Draw a second diagonal with the same origin (x-intercept) as the "normal" line, but a steeper slope.
  • Label this line "noncompetitive inhibitor".
  • Notice that Km does not change in the presence of a noncompetitive inhibitor.
    • This is because the noncompetitive inhibitor interferes with the number of active enzymes, not the affinity of the enzyme for the substrate.
  • Next, label the new 1/Vmax as 1/Vmax(noncomp) on the y-intercept.
    • In the presence of a noncompetitive inhibitor, all active sites are bound by substrate at a lower substrate concentration, which means that 1/Vmax is higher (because Vmax is lowered).
    • Even if the substrate concentration increases, the Vmax will not increase.
  • As a clinical correlate, angiotensin-converting enzyme inhibitors (ACE Inhibitors) are blood pressure lowering agents that noncompetitively inhibit angiotensin-converting enzyme to prevent the formation of angiotensin, which acts on the kidneys to increase blood pressure.