MCAT Biochemistry Review

Chapter 2: Enzymes

Conclusion

Our current chapter focused on the way in which cells are able to carry out the reactions necessary for life. We began with a discussion of the types of enzymes that you are likely to encounter on Test Day before reviewing thermodynamics and kinetics in relation to enzymes, which are biological catalysts. We went on to discuss the analysis of kinetic data with two different types of graphs, and talked about cooperativity. Because catalysts are generally most active in their native environment, we considered the impact of temperature, pH, and salinity on their activity. All of these are likely to appear on Test Day.

Enzymes need to be regulated; we analyzed the basics of feedback mechanisms. We talked about inhibitors of enzymes, which may be reversible or irreversible. The difference between the types of reversible inhibition is a key Test Day concept. Finally, we discussed changes in enzyme activity that may include allosteric activation, covalent modification, or cleavage of inactive zymogens. Let's move on now to discuss the nonenzymatic functions of proteins. You will notice many parallels between the new material and the concepts described in this chapter, like binding affinity. By the end of the next chapter, you'll be ready to face any protein question the MCAT can throw at you!

Concept Summary

Enzymes as Biological Catalysts

·        Enzymes are biological catalysts that are unchanged by the reactions they catalyze and are reusable.

·        Each enzyme catalyzes a single reaction or type of reaction with high specificity.

o   Oxidoreductases catalyze oxidation–reduction reactions that involve the transfer of electrons.

o   Transferases move a functional group from one molecule to another molecule.

o   Hydrolases catalyze cleavage with the addition of water.

o   Lyases catalyze cleavage without the addition of water and without the transfer of electrons. The reverse reaction (synthesis) is often more important biologically.

o   Isomerases catalyze the interconversion of isomers, including both constitutional isomers and stereoisomers.

o   Ligases are responsible for joining two large biomolecules, often of the same type.

·        Exergonic reactions release energy; ΔG is negative.

·        Enzymes lower the activation energy necessary for biological reactions.

·        Enzymes do not alter the free energy (ΔG) or enthalpy (ΔH) change that accompanies the reaction nor the final equilibrium position; rather, they change the rate (kinetics) at which equilibrium is reached.

Mechanisms of Enzyme Activity

·        Enzymes act by stabilizing the transition state, providing a favorable microenvironment, or bonding with the substrate molecules.

·        Enzymes have an active site, which is the site of catalysis.

·        Binding to the active site is explained by the lock and key theory or the induced fit model.

o   The lock and key theory hypothesizes that the enzyme and substrate are exactly complementary.

o   The induced fit model hypothesizes that the enzyme and substrate undergo conformational changes to interact fully.

·        Some enzymes require metal cation cofactors or small organic coenzymes to be active.

Enzyme Kinetics

·        Enzymes experience saturation kinetics: as substrate concentration increases, the reaction rate does as well until a maximum value is reached.

·        Michaelis–Menten and Lineweaver–Burk plots represent this relationship as a hyperbola and line, respectively.

·        Enzymes can be compared on the basis of their Km and vmax values.

·        Cooperative enzymes display a sigmoidal curve because of the change in activity with substrate binding.

Effects of Local Conditions on Enzyme Activity

·        Temperature and pH affect an enzyme's activity in vivo; changes in temperature and pH can result in denaturing of the enzyme and loss of activity due to loss of secondary, tertiary, or, if present, quaternary structure.

·        In vitro, salinity can impact the action of enzymes.

Regulation of Enzyme Activity

·        Enzyme pathways are highly regulated and subject to inhibition and activation.

·        Feedback inhibition is a regulatory mechanism whereby the catalytic activity of an enzyme is inhibited by the presence of high levels of a product later in the same pathway.

·        Reversible inhibition is characterized by the ability to replace the inhibitor with a compound of greater affinity or to remove it using mild laboratory treatment.

o   Competitive inhibition results when the inhibitor is similar to the substrate and binds at the active site. Competitive inhibition can be overcome by adding more substrate. vmax is unchanged, Km increases.

o   Noncompetitive inhibition results when the inhibitor binds with equal affinity to the enzyme and the enzyme–substrate complex. vmax is decreased, Km is unchanged.

o   Mixed inhibition results when the inhibitor binds with unequal affinity to the enzyme and the enzyme–substrate complex. vmax is decreased, Km is increased or decreased depending on if the inhibitor has higher affinity for the enzyme or enzyme–substrate complex.

o   Uncompetitive inhibition results when the inhibitor binds only with the enzyme–substrate complex. Km and vmax both decrease.

·        Irreversible inhibition alters the enzyme in such a way that the active site is unavailable for a prolonged duration or permanently; new enzyme molecules must be synthesized for the reaction to occur again.

·        Regulatory enzymes can experience activation as well as inhibition.

o   Allosteric sites can be occupied by activators which increase either affinity or enzymatic turnover.

o   Phosphorylation (covalent modification with phosphate) or glycosylation (covalent modification with carbohydrate) can alter the activity or selectivity of enzymes.

o   Zymogens are secreted in an inactive form and are activated by cleavage.

Answers to Concept Checks

·        2.1

1.    Catalysts are characterized by two main properties: they reduce the activation energy of a reaction, thus speeding up the reaction, and they are not used up in the course of the reaction. Enzymes improve the environment in which a particular reaction takes place, which lowers its activation energy. They are also regenerated at the end of the reaction to their original form.

2.    Enzyme specificity refers to the idea that a given enzyme will only catalyze a given reaction or type of reaction. For example, serine/threonine-specific protein kinases will only place a phosphate group onto the hydroxyl group of a serine or threonine residue.

3.     

Name

Function

Ligase

Addition or synthesis reactions, generally between large molecules; often require ATP

Isomerase

Rearrangement of bonds within a compound

Lyase

Cleavage of a single molecule into two products, or synthesis of small organic molecules

Hydrolase

Breaking of a compound into two molecules using the addition of water

Oxidoreductase

Oxidation–reduction reactions (transferring electrons)

Transferase

Movement of a functional group from one molecule to another

4.    Enzymes have no effect on the overall thermodynamics of the reaction; they have no effect on the ΔG or ΔH of the reaction, although they do lower the energy of the transition state, thus lowering the activation energy. However, enzymes have a profound effect on the kinetics of a reaction. By lowering activation energy, equilibrium can be achieved faster (although the equilibrium position does not change).

·        2.2

1.     

Lock and Key

Induced Fit

·        Active site of enzyme fits exactly around substrate

·        No alterations to tertiary or quaternary structure of enzyme

·        Less accurate model

·        Active site of enzyme molds itself around substrate only when substrate is present

·        Tertiary and quaternary structure is modified for enzyme to function

·        More accurate model

2.    Cofactors and coenzymes both act as activators of enzymes. Cofactors tend to be inorganic (minerals), while coenzymes tend to be small organic compounds (vitamins). In both cases, these regulators induce a conformational change in the enzyme that promotes its activity. Tightly bound cofactors or coenzymes that are necessary for enzyme function are termed prosthetic groups.

·        2.3

1.    Increasing [S] has different effects, depending on how much substrate is present to begin with. When the substrate concentration is low, an increase in [S] causes a proportional increase in enzyme activity. At high [S], however, when the enzyme is saturated, increasing [S] has no effect on activity because vmax has already been attained. 
Increasing [E] will always increase vmax, regardless of the starting concentration of enzyme.

2.    Both the Michaelis–Menten and Lineweaver–Burk relationships account for the values of Km and vmax under various conditions. They both provide simple graphical interpretations of these two variables and are derived from the Michaelis–Menten equation. However, the axes of these graphs and visual representation of this information is different between the two. The Michaelis–Menten plot is v vs. [S], which creates a hyperbolic curve for monomeric enzymes. The Lineweaver–Burk plot, on the other hand, is  vs , which creates a straight line.

3.    Km is a measure of an enzyme's affinity for its substrate, and is defined as the substrate concentration when an enzyme is functioning at half of its maximal velocity. As Km increases, an enzyme's affinity for its substrate decreases.

4.    The x-intercept represents  ; the y-intercept represents  .

5.    Cooperativity refers to the interactions between subunits in a multisubunit enzyme or protein. The binding of substrate to one subunit induces a change in the other subunits from the T (tense) state to the R (relaxed) state, which encourages binding of substrate to the other subunits. In the reverse direction, the unbinding of substrate from one subunit induces a change from R to T in the remaining subunits, promoting unbinding of substrate from the remaining subunits.

·        2.4

1.    As temperature increases, enzyme activity generally increases (doubling approximately every 10°C). Above body temperature, however, enzyme activity quickly drops off as the enzyme denatures. 
Enzymes are maximally active within a small pH range; outside of this range, activity drops quickly with changes in pH as the ionization of the active site changes and the protein is denatured. 
Changes in salinity can disrupt bonds within an enzyme, causing disruption of tertiary and quaternary structure, which leads to loss of enzyme function.

2.    Ideal temperature: 37°C = 98.6°F = 310 K

Ideal pH for most enzymes is 7.4; for gastric enzymes, around 2; for pancreatic enzymes, around 8.5.

·        2.5

1.    Feedback inhibition refers to the product of an enzymatic pathway turning off enzymes further back in that same pathway. This helps maintain homeostasis: as product levels rise, the pathway creating that product is appropriately downregulated.

2.    The four types of inhibitors are: competitive, noncompetitive, mixed, and uncompetitive.

3.    Irreversible inhibition refers to the prolonged or permanent inactivation of an enzyme, such that it cannot be easily renatured to gain function.

4.    Examples of transient modifications include allosteric activation or inhibition. Examples of covalent modifications include phosphorylation and glycosylation.

5.    Zymogens are precursors of an active enzyme. It is critical that certain enzymes (like the digestive enzymes of the pancreas) remain inactive until arriving at their target site.

Equations to Remember

(2.1) Michaelis–Menten rates

(2.2) Michaelis–Menten equation

Shared Concepts

·        Biochemistry Chapter 1

o   Amino Acids, Peptides, and Proteins

·        Biochemistry Chapter 12

o   Bioenergetics and Regulation of Metabolism

·        Biology Chapter 9

o   The Digestive System

·        General Chemistry Chapter 5

o   Chemical Kinetics

·        General Chemistry Chapter 7

o   Thermochemistry

·        General Chemistry Chapter 11

o   Oxidation–Reduction Reactions