Induced Fit and Enzyme Function

For many years, scientists thought that enzyme-substrate binding took place in a simple “lock-and-key” fashion. This model asserted that the enzyme and substrate fit together perfectly in one instantaneous step. However, current research supports a more refined view scientists call induced fit ((Figure)). This model expands upon the lock-and-key model by describing a more dynamic interaction between enzyme and substrate. As the enzyme and substrate come together, their interaction causes a mild shift in the enzyme’s structure that confirms an ideal binding arrangement between the enzyme and the substrate’s transition state. This ideal binding maximizes the enzyme’s ability to catalyze its reaction.

When an enzyme binds its substrate, it forms an enzyme-substrate complex. This complex lowers the reaction’s activation energy and promotes its rapid progression in one of many ways. On a basic level, enzymes promote chemical reactions that involve more than one substrate by bringing the substrates together in an optimal orientation. The appropriate region (atoms and bonds) of one molecule is juxtaposed to the other molecule’s appropriate region with which it must react. Another way in which enzymes promote substrate reaction is by creating an optimal environment within the active site for the reaction to occur. Certain chemical reactions might proceed best in a slightly acidic or non-polar environment. The chemical properties that emerge from the particular arrangement of amino acid residues within an active site create the perfect environment for an enzyme’s specific substrates to react.

You have learned that the activation energy required for many reactions includes the energy involved in manipulating or slightly contorting chemical bonds so that they can easily break and allow others to reform. Enzymatic action can aid this process. The enzyme-substrate complex can lower the activation energy by contorting substrate molecules in such a way as to facilitate bond-breaking, helping to reach the transition state. Finally, enzymes can also lower activation energies by taking part in the chemical reaction itself. The amino acid residues can provide certain ions or chemical groups that actually form covalent bonds with substrate molecules as a necessary step of the reaction process. In these cases, it is important to remember that the enzyme will always return to its original state at the reaction’s completion. One of enzymes’ hallmark properties is that they remain ultimately unchanged by the reactions they catalyze. After an enzyme catalyzes a reaction, it releases its product(s).

Figure 2. According to the induced-fit model, both enzyme and substrate undergo dynamic conformational changes upon binding. The enzyme contorts the substrate into its transition state, thereby increasing the reaction’s rate.

Molecular Regulation of Enzymes

Enzymes can be regulated in ways that either promote or reduce their activity. There are many different kinds of molecules that inhibit or promote enzyme function, and various mechanisms exist for doing so. For example, in some cases of enzyme inhibition, an inhibitor molecule is similar enough to a substrate that it can bind to the active site and simply block the substrate from binding. When this happens, the enzyme is inhibited through competitive inhibition, because an inhibitor molecule competes with the substrate for active site binding ((Figure)). Alternatively, in noncompetitive inhibition, an inhibitor molecule binds to the enzyme in a location other than an allosteric site, a binding site away from the active site, and still manages to block substrate binding to the active site.

Figure 3. Competitive and noncompetitive inhibition affect the reaction’s rate differently. Competitive inhibitors affect the initial rate but do not affect the maximal rate; whereas, noncompetitive inhibitors affect the maximal rate.

Some inhibitor molecules bind to enzymes in a location where their binding induces a conformational change that reduces the enzyme’s affinity for its substrate. This type of inhibition is an allosteric inhibition ((Figure)). More than one polypeptide comprise most allosterically regulated enzymes, meaning that they have more than one protein subunit. When an allosteric inhibitor binds to an enzyme, all active sites on the protein subunits change slightly such that they bind their substrates with less efficiency. There are allosteric activators as well as inhibitors. Allosteric activators bind to locations on an enzyme away from the active site, inducing a conformational change that increases the affinity of the enzyme’s active site(s) for its substrate(s).

Figure 4. Allosteric inhibitors modify the enzyme’s active site so that substrate binding is reduced or prevented. In contrast, allosteric activators modify the enzyme’s active site so that the affinity for the substrate increases.

Everyday Connection

Figure 5. Have you ever wondered how pharmaceutical drugs are developed? (credit: Deborah Austin)

 

Drug Discovery by Looking for Inhibitors of Key Enzymes in Specific Pathways

Enzymes are key components of metabolic pathways. Understanding how enzymes work and how they can be regulated is a key principle behind developing many pharmaceutical drugs ((Figure)) on the market today. Biologists working in this field collaborate with other scientists, usually chemists, to design drugs.

Consider statins for example—which is a class of drugs that reduces cholesterol levels. These compounds are essentially inhibitors of the enzyme HMG-CoA reductase. HMG-CoA reductase is the enzyme that synthesizes cholesterol from lipids in the body. By inhibiting this enzyme, the drug reduces cholesterol levels synthesized in the body. Similarly, acetaminophen, popularly marketed under the brand name Tylenol, is an inhibitor of the enzyme cyclooxygenase. While it is effective in providing relief from fever and inflammation (pain), scientists still do not completely understand its mechanism of action.

How are drugs developed? One of the first challenges in drug development is identifying the specific molecule that the drug is intended to target. In the case of statins, HMG-CoA reductase is the drug target. Researchers identify targets through painstaking research in the laboratory. Identifying the target alone is not sufficient. Scientists also need to know how the target acts inside the cell and which reactions go awry in the case of disease. Once researchers identify the target and the pathway, then the actual drug design process begins. During this stage, chemists and biologists work together to design and synthesize molecules that can either block or activate a particular reaction. However, this is only the beginning: both if and when a drug prototype is successful in performing its function, then it must undergo many tests from in vitro experiments to clinical trials before it can obtain FDA approval to be on the market.

Many enzymes don’t work optimally, or even at all, unless bound to other specific non-protein helper molecules, either temporarily through ionic or hydrogen bonds or permanently through stronger covalent bonds. Two types of helper molecules are cofactors and coenzymes. Binding to these molecules promotes optimal conformation and function for their respective enzymes. Cofactors are inorganic ions such as iron (Fe++) and magnesium (Mg++). One example of an enzyme that requires a metal ion as a cofactor is the enzyme that builds DNA molecules, DNA polymerase, which requires a bound zinc ion (Zn++) to function. Coenzymes are organic helper molecules, with a basic atomic structure comprised of carbon and hydrogen, which are required for enzyme action. The most common sources of coenzymes are dietary vitamins ((Figure)). Some vitamins are precursors to coenzymes and others act directly as coenzymes. Vitamin C is a coenzyme for multiple enzymes that take part in building the important connective tissue component, collagen. An important step in breaking down glucose to yield energy is catalysis by a multi-enzyme complex scientists call pyruvate dehydrogenase. Pyruvate dehydrogenase is a complex of several enzymes that actually requires one cofactor (a magnesium ion) and five different organic coenzymes to catalyze its specific chemical reaction. Therefore, enzyme function is, in part, regulated by an abundance of various cofactors and coenzymes, which the diets of most organisms supply.

Figure 6. Vitamins are important coenzymes or precursors of coenzymes, and are required for enzymes to function properly. Multivitamin capsules usually contain mixtures of all the vitamins at different percentages.

Enzyme Compartmentalization

In eukaryotic cells, molecules such as enzymes are usually compartmentalized into different organelles. This allows for yet another level of regulation of enzyme activity. Enzymes required only for certain cellular processes are sometimes housed separately along with their substrates, allowing for more efficient chemical reactions. Examples of this sort of enzyme regulation based on location and proximity include the enzymes involved in the latter stages of cellular respiration, which take place exclusively in the mitochondria, and the enzymes involved in digesting cellular debris and foreign materials, located within lysosomes.

Feedback Inhibition in Metabolic Pathways

Molecules can regulate enzyme function in many ways. However, a major question remains: What are these molecules and from where do they come? Some are cofactors and coenzymes, ions, and organic molecules, as you have learned. What other molecules in the cell provide enzymatic regulation, such as allosteric modulation, and competitive and noncompetitive inhibition? The answer is that a wide variety of molecules can perform these roles. Some include pharmaceutical and non-pharmaceutical drugs, toxins, and poisons from the environment. Perhaps the most relevant sources of enzyme regulatory molecules, with respect to cellular metabolism, are cellular metabolic reaction products themselves. In a most efficient and elegant way, cells have evolved to use their own reactions’ products for feedback inhibition of enzyme activity. Feedback inhibition involves using a reaction product to regulate its own further production ((Figure)). The cell responds to the abundance of specific products by slowing down production during anabolic or catabolic reactions. Such reaction products may inhibit the enzymes that catalyzed their production through the mechanisms that we described above.

Figure 7. Metabolic pathways are a series of reactions that multiple enzymes catalyze. Feedback inhibition, where the pathway’s end product inhibits an upstream step, is an important regulatory mechanism in cells.

Producing both amino acids and nucleotides is controlled through feedback inhibition. Additionally, ATP is an allosteric regulator of some of the enzymes involved in sugar’s catabolic breakdown, the process that produces ATP. In this way, when ATP is abundant, the cell can prevent its further production. Remember that ATP is an unstable molecule that can spontaneously dissociate into ADP. If too much ATP were present in a cell, much of it would go to waste. Alternatively, ADP serves as a positive allosteric regulator (an allosteric activator) for some of the same enzymes that ATP inhibits. Thus, when relative ATP levels are high compared to ATP, the cell is triggered to produce more ATP through sugar catabolism.