Enzymes

Enzyme Structure

Enzymes are proteins that work as catalysts, or molecules that speed up biochemical reactions.

Cells perform many, many biochemical reactions, both to build and break down materials. If it were not for proteins called enzymes that speed up reactions, these processes would take too long for the cell to survive. An enzyme is a substance, usually a protein, that speeds up a biological reaction without being consumed in the reaction. Enzymes participate in reactions without being altered themselves so that a single enzyme may interact with a large number of target molecules. Some processes require more than one enzyme. Glycolysis, the chemical pathway that allows cells to obtain energy from glucose, uses 10 different enzymes at different steps.

An enzyme is a protein that binds with a specific substrate, or target molecule. It serves as a catalyst, speeding up the chemical reactions in cells. A living cell carries out hundreds of different metabolic pathways. A metabolic pathway is a series of chemical reactions that occur in a living organism and may be facilitated by enzymes. The products of one set of enzymatic reaction serve as the reactants for a different enzymatic reaction. Each step of the pathway has an associated enzyme, and each enzyme has evolved structures specifically suited to the efficient performance of its job. There are thousands of different enzymes in cells. Each type of enzyme catalyzes a particular reaction involving a specific substrate.

The tertiary structure of an enzyme, which is a structure of a protein produced by interactions between the R groups of the amino acids in the chain and the environment around them, frequently creates an active site that is specific to the enzyme's substrate. Because the active site is formed by the tertiary folding of the protein, the amino acids important for the active site may be distant from one another in the enzyme's primary structure. This active site is the portion of the enzyme that binds to the substrate by a variety of noncovalent interactions and catalyzes the reaction. The enzyme holds the substrate in position for the reaction to proceed. Some enzymes may catalyze a reaction between two substrates. In these cases, the enzyme positions the two substrates so that their interaction is enabled. It is also possible that the active site is created by the interaction in the quaternary state.

The specificity of an enzyme and its substrate led scientists to characterize their structure as a lock and key, with the substrate fitting precisely into the enzyme at the active site. However, many enzyme-substrate interactions are better described as an induced fit, meaning that both enzyme and substrate change shape upon binding. The distortion of bonds caused by this shape change assists in the process of catalysis, the act of a catalyst, such as an enzyme, speeding up a biological reaction.

Enzyme with Bound Substrate

The structure of the enzyme binds its substrate, peptidoglycan, in a cleft between two globular protein chains of its tertiary structure. The side chains of the region help to position the substrate correctly. The side chains near the active site aid in catalysis.

How Enzymes Affect Activation Energy

Enzymes lower the activation energy needed to begin a specific chemical reaction.

Chemical reactions proceed from reactants to products. The speed of a reaction depends on the activation energy, or the amount of energy required for the reaction to initiate. In enzyme-catalyzed reactions, the reactant is the substrate molecule.

When the enzyme-substrate complex forms, the activation energy of the reaction decreases. This can happen in a number of ways. Chemical bonds in the substrate are strained by the induced fit of the substrate to the enzyme and become easier to break. In situations where the binding of two molecules is catalyzed by an enzyme, the two substrates are brought into alignment to promote bonding. Some chemically active side chains of the enzyme itself may participate directly in the reaction, forming intermediate states with the substrate. Enzymes may also bind additional substances called cofactors, which are small molecules or metal ions that aid in catalysis. These changes allow enzyme-catalyzed reactions to proceed rapidly.

{\text {Enzyme}}+{\text {Substrate}}\Leftrightarrow{\text{Enzyme}} - {\text {Substrate Complex}}\Leftrightarrow {\text {Enzyme}} + {\text {Product}}

The role of the enzyme carbonic anhydrase in converting carbonic acid to water and carbon dioxide (and vice versa) illustrates the importance of enzyme-catalyzed reactions for living things. Animals, including humans, take in oxygen from the air by breathing. Within the cells, the mitochondria use oxygen for metabolic processes and produce carbon dioxide as a waste product. The carbon dioxide must then be removed from the body. It passes out of the cells of the tissue where it was generated and into the blood. Carbonic anhydrase in the red blood cells quickly converts the carbon dioxide into carbonic acid (some conversions happen much more slowly in the blood plasma as well). The red blood cells transport the carbonic acid (as the bicarbonate ion) to the lungs, where carbonic anhydrase catalyzes the reverse process, this time converting the carbonic acid to carbon dioxide gas, which is then exhaled. Carbonic anhydrase is essential to making this process happen on the timescale needed by an organism. Both reactions happen slowly in the absence of the enzyme but occur up to a million times faster when catalyzed by carbonic anhydrase. Because a buildup of carbon dioxide is unhealthy for animals, including humans, carbonic anhydrase is critical to survival. Carbonic anhydrase is also found in plants, where it is important in converting stored bicarbonate ions to carbon dioxide for photosynthesis. It is also found in algae and bacteria. There is a wide variety of sequences and structures among the forms of carbonic anhydrase found in these organisms, which is evidence for convergent evolution driven by the importance of this enzyme.

Enzymes Lower the Activation Energy of Reactions

An enzyme increases the rate of a reaction by lowering the activation energy required for the reaction to proceed. A reaction catalyzed by an enzyme may be millions of times faster than the same reaction without the enzyme.

Active Sites

The shape and surface charges of an enzyme play a critical role in effecting enzymatic reactions by creating active sites where the substrate binds and the reaction occurs.

Enzymes have two primary ways of creating an active site specific for a particular substrate. These are (1) the shape of the active site, which is the place on an enzyme where the substrate binds and the reaction occurs, and (2) the varying surface charge of the enzyme at locations both within and outside the active site. The 20 different R groups, or side chains, of amino acids offer a wide range of sizes, shapes, mobility, and levels of electrostatic charge. The tertiary structure of the protein evolves so that the side chains in the active site are precisely configured to selectively bind the substrate and catalyze the reaction. Even water molecules are often excluded from the active site unless they are involved in catalyzing the reaction.

The specificity of the active site chemistry occurs in different ways. A few charged side chains may be clustered so that they coordinate (bind to) a metal ion that helps the reaction proceed, as happens in carbonic anhydrase. Alternatively, the size, shape, and electrostatic nature (hydrophobic, polar, or charged) of the reaction site may select a specific substrate, as occurs with a family of enzymes called serine proteases.

Beyond the active site, the entire surface of the enzyme contains patches of positive and negative charge that can contribute to substrate binding. This is referred to as the electrostatic potential map of the protein surface. These larger-scale charged areas are thought to be important in guiding the approaching substrate to bind to the active site. The overall shape of the enzyme contributes as well. If the tertiary structure of the enzyme provides a shape (sometimes called a binding cleft or binding pocket) that is complementary to the substrate, this promotes tighter binding.

The importance of the specific side chains involved in these interactions has been demonstrated experimentally. Researchers have created versions of enzymes in which specific amino acids in the active site were changed. When the replaced side chain was involved in catalyzing the reaction, this resulted in a change (typically a drop) in the reaction rate. The substrate may not have been bound as tightly, or it may have been less optimally oriented, resulting in less efficient catalysis.

Controlling Enzyme Activity

Cells closely regulate enzyme activity through feedback inhibition, allosteric regulation, and competitive and noncompetitive inhibition.

Cells need ways to control the activity of enzymes, either to avoid overproducing the product of a reaction or to trigger production in response to environmental conditions. One means of control is through feedback inhibition, in which the product of the reaction binds to the enzyme to change the conformation of the active site, thus deactivating it. As the concentration of product builds up, the enzymatic activity decreases. Allosteric regulation is a type of enzyme regulation in which the location at which the regulatory molecule binds is not the same as the active site. Feedback inhibition is a form of allosteric regulation, where the product does not bind to the active site, but to a different location on the enzyme. The term "allosteric regulation" comes from the words allo- meaning "other" and -steric meaning "site." As a result of the product binding to this other location, the enzyme changes its conformation, thereby halting its ability to continue catalyzing reactions.

Allosteric modulation refers to the ability of a substance to indirectly influence the efficacy or binding ability of the enzyme’s primary ligand. A ligand is a substance that forms a complex with a larger biomolecule for a specific purpose. Positive allosteric modulators enhance the effects of the primary ligand. Negative allosteric modulators decrease the effects of the primary ligand. Silent allosteric modulators occupy the allosteric site and are functionally neutral.

Another means of enzyme regulation is through the action of an inhibitor, or a molecule (other than the products of the reaction) that decreases the rate of reaction. Some inhibitors occur naturally, while others are human-designed molecules found in drugs and pesticides. A competitive inhibitor binds to the active site of the enzyme, denying substrate molecules access to the enzyme. Because they are not covalently bound to the enzyme, competitive inhibition can be overcome by increasing the concentration of substrate.

Competitive Enzyme Inhibition

In competitive enzyme inhibition, an inhibiting agent temporarily or transiently prevents the enzyme from binding to the substrate.

Noncompetitive inhibition, on the other hand, is the result of a type of allosteric inhibitor. It does not compete with the substrate for binding to the active site, but binds elsewhere and changes the shape of the enzyme so that it can no longer bind to the substrate. Feedback inhibition and noncompetitive inhibition are both forms of allosteric inhibition, with the end result being a decrease in the amount of enzyme available for chemical binding.

Some enzymes are controlled by other enzymes, or through covalent modification, which is a process through which enzymes alter proteins. Such changes can occur to single amino acids or complete amino acid chains. The modifying enzyme may add a chemical group, often a phosphate or a methyl group, to the enzyme being regulated. Covalent modifications may either activate enzymes or inhibit them.

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