Proteins and Enzymes

Assessment Statements

7.5.1 Explain the four levels of protein structure, indicating the significance of each level.

7.5.2 Outline the difference between fibrous and globular proteins, with reference to two examples of each protein type.

7.5.3 Explain the significance of polar and non-polar amino acids.

7.5.4 State the four functions of proteins giving a named example of each.

7.6.1 State that metabolic pathways consist of chains and cycles or enzyme-catalyzed reactions.

7.6.2 Describe the induced fit model

7.6.3 Explain that enzymes lower the activation energy of the chemical reactions that they catalyze

7.6.4 Explain the difference between competitive and non-competitive inhibition, with reference to one example of each.

7.6.5 Explain the control of metabolic pathways by end product inhibition, including the role of allosteric sites.

So, the protein has been made using the translation process. What is it used for?

Some proteins have amino acids that are acidic, basic, have hydrophobic properties and have hydrophilic properties. There are proteins that perform structural tasks, proteins that store amino acids and some that have receptor functions so cells can respond to chemical signals. In order to do this, they need to combine into many forms or structures. The function of a protein is very closely related to its structure. See worksheet on Amino Acids.

Protein Structure (See worksheet – Proteins)

Proteins have four levels of organization:

1. Primary Organization - this is the unique sequence of amino acids held together by peptide bonds. The order or sequence was determined by the nucleotide base sequence on the DNA. Every organism has its own DNA, and therefore, every organism has its own unique proteins. The significance of the primary structure is the sequence determines the higher levels of the protein. Changing one amino acid may completely alter the structure and function of a protein. Ex. Sickle cell disease.

2. Secondary Organization – this is created when the hydrogen bonds form between the oxygen from the carboxyl group from one amino acid and the hydrogen from the amino group of another. It does not include side chains, the R groups. The significance of the structure is that, even though the hydrogen bonds are weak, several of them in a row provide strength. The most common configurations are the helix and the sheet.

3. Tertiary Organization – this is when the polypeptide chain bends and folds over itself, based upon the interactions among the R groups and the peptide backbone. The result is a definite three dimensional conformation. Interactions that cause tertiary conformations are:

a. Covalent bonds between sulfur atoms to create disulfide bonds or bridges because they are so strong

b. Hydrogen bonds between the side chainsc. Van der Waals interactions among the hydrophobic side chains on the amino acids.

These interactions are strong because many hydrophobic side chains are forced inwards when the hydrophilic side chains interact with water towards the outside of the molecule.

d. Ionic bonds between positively and negatively charged side chains.Tertiary Organization is significant, because it determines the specificity of the proteins known as enzymes.

4. Quaternary Organization – this is unique in that is involves multiple polypeptide chains which combine to form a single structure. Not all proteins have this organization. One example is haemoglobin, which is four protein chains held around a haem group, in which an atom of iron occurs. These proteins are called conjugated proteins. This is significant because the structure is very specialized in terms of its actions.

Fibrous and globular proteins are examples of tertiary organization. Fibrous proteins are those that are polypeptide chains in a long narrow shape. Examples are collagen, which plays a role in the connective tissue of humans and actin. Globular proteins are more three dimensional in their shape. Examples are haemoglobin and insulin. Use the worksheet to add to these notes, as to the differences between them.

Polar and Non-polar Amino Acids

Amino acids are grouped according to their side chains.

Amino acids with non-polar side chains are hydrophobic. They are found in the regions of proteins that are linked to the hydrophobic area of the cell membrane. This means they can remain embedded in the membrane and attract non-polar substances, like fats/lipids. They will also repel water, which is polar.

Amino acids with polar side chains are hydrophilic. They are found in regions of proteins that are exposed to water. Membrane proteins include polar amino acids towards the interior and exterior of the membrane, to create polar channels. Polar substances can move through these channels.

Polar and non-polar amino acids are important in determining the specificity of an enzyme. Each enzyme has a region called the active site. Only specific substrates can combine with particular active sites. Combination is possible when fitting occurs. The “fitting” involves the general shapes and polar properties of the substrate and of the amino acids exposed at the active site.

The Functions of Proteins

Proteins can be membranous and non-membranous.

Membrane Proteins, as a review from last year, are responsible for:

1. Hormone Bonding Sites2. Enzyme Receptor Sites3. Electron Carriers4. Channels for Passive Transport5. Pumps for Active Transport

Non-membrane Proteins are responsible for:

1. Structural Support – Collagen and Keratin (hair)2. Transport – Haemoglobin3. Movement – Myosin and actin4. Defense – Immunoglobulin or antibodies

Others are chemical messengers, or hormones, like insulin and ADH, while others are biological catalysts, or enzymes, like pepsin. Others are food stores, like casein in milk and pigments, like opsin in our eyes and some organisms use them as toxins, like snake venom.

Enzymes

Metabolism is the sum of all the chemical reactions in your body. The reactions to build molecules are called anabolic, and those that break down molecules, are called catabolic. Anabolic reactions require energy and Catabolic reactions release energy.

Almost all metabolic reactions in organisms are catalyzed by enzymes. Many of the reactions occur in metabolic or biochemical pathways that occur in a chain or a cycle. A pathway may be like so:

Enzyme 1 Enzyme 2Substrate A Substrate B Final Product

Most metabolic pathways are carried out in designated compartments of the cell, where the necessary enzymes are clustered and isolated. The enzymes are determined by the cell’s genetic make-up.

The Induced Fit Model (See worksheet on Enzymes)

Last year, you learned about the “lock and key model” of enzyme function. Enzymes are complex proteins that have unique areas, such as the active sites, where it binds to a particular substrate. The lock and key model worked, but now we know more about enzymes.

Now we see that enzymes undergo changes in their conformation, when substrates combine with their active sites. This is called the Induced Fit Model. The binding site changes to fit and accept the substrate. The enzyme then can weaken bonds and reducing the activation energy of the reaction. The process occurs faster. This would explain why some enzymes have such a broad specificity, such as proteases. Therefore, several different but similar substrates could bind to the same enzyme.

A good example is a hand and a glove, the hand being the substrate and the glove being the enzyme. The glove looks like a hand, but only when the hand is placed in the glove, does the glove actually take the shape of the hand. Also, several different hands can fit into the same glove.

The changes that occur in conformation are due to the changes in the R-groups of the amino acids at the active site of the enzyme as they interact with the substrate or substrates.

Mechanism of Enzyme Action

1. The surface of the substrate contacts the active site of the enzyme2. The enzyme changes shape to accommodate the substrate.3. A temporary complex, called the enzyme-substrate complex, or activated complex, forms.4. Activation energy is lowered and the substrate is altered by the rearrangement of existing atoms5. The transformed substrate – product – is released from the active site.6. The unchanged enzyme is then free to combine with other substrate molecules.

Enzyme action can be summarized by:

E + S ES E + P

Where E is the enzyme, S is the substrate, ES is the complex, and P is the product.

Enzymes work by lowering the activation energy of a reaction. Activation Energy, Ae, is the energy necessary to destabilize the existing chemical bonds in a reaction. The reaction occurs faster because the amount of energy needed for the reaction to occur is less. Think about Ae as a wall. You need so much energy to get over the wall, to get to the other side (products). If you can lower the wall, the amount of energy you need is less. This is due to the fact that the active site can weaken the bonds, rearrange them and form the new product.

****Even though the enzymes lower Ae of a particular reaction, they do not alter the proportion of reactants to products.

Read and do the Enzyme Worksheet.

Inhibition and Enzymes

The effects of pH, temperature and substrate concentration were discussed last year. Different molecules have effects on the active sites of enzymes. If a molecule affects the active site in some way, the activity of the enzyme may be altered. This is called inhibition. There are two types: Competitive and Non-Competitive.

Competitive Inhibition

In competitive inhibition, a molecule, called a competitive inhibitor, competes directly for the active site of the enzyme. The result is that the substrate then has fewer encounters with the active site and the chemical reaction rate is decreased. The competitive inhibitor must have a similar structure to the substrate to be able to function in this way.

Competitive inhibition may be reversible or irreversible. Reversible competitive inhibition may be overcome by increasing the substrate concentration. When this happens, there are more substrate molecules to bind to the active sites as they become available, and the reaction my proceed more rapidly.

An example is the use of sulfanilamide (a sulfa drug) to kill the bacteria during an infection. Folic acid is essential as a coenzyme to bacteria. It is produced in bacterial cells by enzyme action on para-aminobenzoic acid (PABA). The sulfanilamide competes with the PABA and blocks the enzyme. Because human cells do not use PABA to produce folic acid, they are unaffected by the drug.

Another example is the reaction between carbon dioxide and the acceptor molecule in photosynthesis, called ribulose bisphosphate carboxylase . When there is oxygen in the chloroplasts, the reaction between the two is inhibited, and photosynthesis slows.

Non-competitive Inhibition

None competitive inhibition involves an inhibitor that does not compete for the enzyme’s active site. The inhibitor interacts with another site on the enzyme. Non-competitive inhibition is also called allosteric inhibition, because the inhibitor binds to a site called the allosteric site. These allosteric enzymes have two non-overlapping binding sites.

Binding at the allosteric site causes a change in the shape of the enzymes active site, making the non-functional.

This type of inhibition may be reversible or irreversible. When the inhibitor concentration is low, an increase in the concentration of substrate increases the enzyme activity. But, since the inhibitor and substrate are not competing for the same site, the substrate cannot prevent the binding of the inhibitor, even when the substrate concentration is high. Some of the enzyme molecules remain inhibited and the maximized activity of the enzyme is lower than when there is no inhibitor.

Examples are opiods that resemble morphine. They inhibit nitric oxide synthase that have signaling roles in the human body. Another example is mercury, which binds to the sulfur groups of component amino acids in enzymes. The shape change causes inhibition of the enzyme.

End Product Inhibition

End product inhibition prevents the cell from wasting chemical resources and energy by making more of a substance than it needs. Many metabolic reactions occur in an assembly line type of process so that a specific end product can be achieved. Each step is catalyzed by a specific enzyme. When the end product is made in a sufficient quantity, the assembly line is shut down. This is usually done by inhibiting the action of the enzyme in the first step of the pathway. This is an example of an allosteric enzyme. The higher concentrations of the end product bind with the active site on the first enzyme, inhibition occurs and the reaction stops. As the existing end product is used up by the cell, the concentrations lower, resulting in fewer bindings with the allosteric site on the first enzyme, and the enzyme is reactivated, resulting in the pathway starting up again.

The bacterium E. coli uses a metabolic pathway to produce the amino acid isoleucine from threonine. It is a 5-step process. If the isoleucine is added to the growth medium of E. coli, it inhibits the first enzyme in the pathway and isoleucine is not synthesized.

The inhibition of the first enzyme is the pathway prevents the build-up of intermediates in the cell. This is a form of negative feedback.