BIO Lecture 5: Enzyme Kinetics and More Inhibition

This lecture is definitely my bread and butter. Therefore I'll try to make this as painless as possible considering the nature of the material ahead.

Enzyme kinetics is simply the study of the rate of formation of products from substrates in the presence of an enzyme. The reaction rate (V denoting velocity) is the amount of product formed per unit time. This depends on the concentration of the substrate [S] and the enzyme. (Before we go any further I want to point out the importance of understanding graphs for this lecture. While I am clearly new at blogging, I have not mastered the art of including images with my lectures. Therefore, please go find a figure in either a biochemistry textbook or online that shows "saturation kinetics" so that you will have a visual to follow along with. In the meantime, I will try my best to find a way to post a drawing or some other type of visual aid.) If there is a small amount of substrate added to the enzyme preparation, enzyme activity appears to increase on the graph almost linearly. However, the enzyme's activity increases less dramatically as more substrate is added. At very high substrate concentrations, enzyme activity appears to level off as it approaches a maximum value (denoted Vmax). At this point the enzyme is said to be saturated. We say that this graph is hyperbolic.

Cooperativity is a different animal. Many multi-subunit enzymes do not behave in the simple kinetic manner described above. In such cases, the binding of substrate to one subunit allosterically increases the affinity of other subunits for the substrate. This is a cooperative case and occurs when the enzyme subunits are structurally linked to each other so that a substrate induced conformational change in one subunit elicits conformational changes in the remaining subunits. EXAMPLE: hemoglobin, when oxygen binding to the heme group in one subunit alters the oxygen affinity of the other subunits. The result is a SIGMOIDAL curve. NOTE: cooperative enzymes must have more than one active site. They are usually multisubunit complexes composed of more than one protein chain held together in a quaternary structure.

INHIBITION: inhibitors reduce enzyme activity through the following mechanisms--competitive inhibition and noncompetitive inhibition.
Competitive: inhibitors compete with substrate for active site (remember that the most effective competitive inhibitors resemble the transition state). NOTE that competitive inhibition can be overcome by adding more substrate; this is because the most substrate we have the more it can out compete the inhibitor. Vmax is not affected but it increases the Km value
Noncompetitive: they bind to an allosteric site, not the active site. Therefore, no matter how much substrate you add, the inhibitor will not be displaced from its site of action. Vmax is diminished therefore.

Now let's touch on the Michaelis constant, Km. Essentially, Km is related to the affinity of the enzyme for the substrate and is the substrate concentration at 1/2 of Vmax. An enzyme with a low Km reaches 1/2 Vmax at very low concentrations because the enzyme has a high affinity for the substrate. An enzyme with a high Km, though, doesn't have a strong affinity for the substrate so it takes a lot more of the substrate to get the enzyme up to 1/2 Vmax. If you haven't done so already, please look at a Lineweaver-Burke plot and a velocity vs. concentration graph for this to make total sense.

One last thing: notice that there is no mention of the Michaelis-Menton equation or the Lineweaver-Burke equation. Questions involving those are usually memorization with plug & chug calculation. Understanding what happens on the graphs is much more intuitive.