AS BIOLOGY - EVERYTHING on enzymes OCR F212

Enzymes are biological catalysts (a substance that speeds up the rate of reaction without being used up in the reaction). They catalyse the two types of metabolic reactions in your body;

ANABOLIC - where small molecules are built up into larger, more complex ones.

CATABOLIC - where larger molecules are broken down into smaller ones.

The way in which enzymes act as a catalyst is by lowering the activation energy of a reaction. All reactions require a certain amount of energy for it to start. Enzymes lower the energy required, which speeds up the rate of reaction. Its the enzyme-substrate complex that lowers the activation energy. Here's one reason;

During a breakdown reaction, the substrate fitting into the active site of an enzyme puts strain on the bonds of the substrate, allowing the molecule to break up more easily.

Their action may be intracellular (within cells) eg. respiration inside cells, or extracellular (outside cell) eg. in digestive system.

Enzymes are usually named by taking the name of the substrate and adding the suffix -ase. This is useful to know if a question asks what type of enzyme is used in a given reaction. The main six broad categories are;

HYDROLASES - Break chemical bonds by the addition of water (hydrolysis). 

OXIDOREDUCTASES - Involved in redox reactions.

TRANSFERASES - Transfer a group of atoms from one molecule to another.

ISOMERASES - Control the conversion of one isomer to another.

LYASES - Break chemical bonds without the addition of water.

LIGASES - Form new chemical bonds using ATP as a source of energy.

PROTEASE (hydrolase) - They hydrolyse proteins to smaller peptides.

ALCOHOL DEHYDROGENASE (oxidoreductase) - They facilitate the interconversion of alcohols to aldehydes or ketones.

METHYLTRANSFERASE (transferase) - Transfers a methyl group from a donor to an acceptor.

PHOSPHOGLUCOMUTASE (isomerase) - They facilitate the interconversion of glucose 1-phosphate and glucose 6-phosphate.

DECARBOXYLASE (lyase) - They add or remove a carboxyl group from organic compounds.

DNA LIGASE (ligase) - repairs single-stranded discontinuities in double stranded DNA molecules.

THIS DOES NOT NEED TO BE KNOWN, IT IS JUST SIMPLY HERE TO HELP YOUR UNDERSTANDING OF THE SIX MAIN GROUPS.

Enzymes are globular proteins and are very specific. They usually only catalyse one reaction due to only one substrate being able to match the shape of the active site. They have three levels of protein structure.

PRIMARY - This refers to the sequence of amino acids in the polypeptide chain. Each enzyme has its own sequence of amino acids that makes it different from any other enzyme. This also determines the tertiary structure.

SECONDARY - This refers to the twisting of the amino acid chain to form the most stable pattern of hydrogen bonding. In globular proteins the most common arrangements are the alpha helix shape or beta pleated sheet.

TERTIARY - This refers to how the secondary structure folds back on itself to form its own complex three dimensional shape. This structure causes the specificity of an enzyme as every enzyme has a different tertiary structure. If the tertiary structure is altered in any way, the shape of the active site will change resulting in denaturation. 

Enzymes only work with the substrate that fits their active site. The lock and key model was an early description to attempt to explain the specificity of enzymes. The theory states that;

Enzyme function depends on the area of the molecule known as the active site. 

The size, shape and chemical nature of the active site corresponds so closely to the substrate molecule that they fit together like a key in a lock.

The substrate and enzyme momentarily bond at the active site to form an enzyme-substrate complex.

The resulting tension or proximity causes the reaction to occur and the product(s) to formed.

The product(s) has a different shape to the substrate and so no longer fits the active site so as a result, it is released.

The enzyme is then free to interact with more substrate.

The lock and key model suggests an exact fit between the active site and substrate.

However more recent x-ray diffraction studies have shown that the shape of an enzyme molecule alone differs from its shape when it combines with the substrate. Significant changes take place in the shape of an enzyme when it binds.

Hence the induced fit theory states;

The binding of the substrate to the active site causes the enzyme to change shape so that the two molecules fit together more snugly.

BOTH OF THE TWO THEORIES DEMONSTRATE THE IMPORTANCE OF THE TERTIARY STRUCTURE OF THE ENZYME MOLECULE TO ITS FUNCTION.

The energy needed to start a chemical reaction in an organism is known as the activation energy. Enzymes lower this activation energy.

Like any chemical reaction, the rate of an enzyme-controlled reaction increases when the temperature increases. This is because more heat provides more kinetic energy for both the enzyme and substrate molecules. This means they both move faster, increasing the chances of a collision. The energy of the collisions also increases, meaning each collision is more likely to result in a reaction. 

But for enzyme-controlled reactions, this temperature increase only continues up to a point after which the rate of reaction declines. This occurs because the heating causes the atoms, that make up the enzyme molecule, to vibrate more. Vigorous enough vibrations will cause the hydrogen bonds that determine the structure of the enzyme to break.

This leads to the loss of the specific shape of the active site and so the enzyme becomes inactive. This loss of structure is PERMANENT. It cannot be reversed by cooling. The enzyme has been denatured ie. it no longer functions.

Most enzymes have an optimum pH at which rate of reaction is fastest eg. pepsin works best at acidic pH 2. As you know, the tertiary structure held together by hydrogen bonds is key to the enzymes function. Small changes in pH can affect the rate of reaction without denaturing the enzyme. But extreme changes can cause the enzyme to become unstable and denatured.

Acidity and alkalinity can affect the site of an enzyme. Free H+ and OH- ions can affect the charges on the amino acid side chains of an enzymes active site. This will affect the hydrogen bonding and so change the 3 dimensional shape of an enzyme and the shape of the active site. Therefore, the substrate will no longer fit the active site and the enzyme loses its activity and the rate of reaction falls.

If the active site becomes flooded with H+ or OH- ions then it can prevent the enzyme and substrate from fitting together. If the enzyme and substrate have the same charges, they repel each other and an enzyme-substrate complex is not formed. 

The higher the substrate concentration, the faster the rate of reaction. This is because there are more substrate molecules which means a collision between and enzyme and substrate is more likely to occur, and so more active sites will be used. 

However, this only occurs up to a point known as 'saturation', whereby there are so many substrate molecules that all the active sites are currently full and so the rate of reaction cannot increase. So the graph (next slide) plateaus off. 

Adding more substrate molecules makes no more difference.

Point A - Volume of substrate molecules is being increased.

Point B - Saturation, all active sites are currently full.

The more enzyme molecules there are, the more likely a collision will occur between a substrate and enzyme molecule. So an increase in concentration of enzyme will result in an increased rate of reaction. If there is an excess of substrate (ie. substrate concentration is not limited) then the rate of reaction and enzyme concentration are said to be directly proportional - when one increases, so does the other (DASHED LINE ON GRAPH)

However, when the amount of substrate is limited, there comes a point whereby there is more enzyme molecules than substrate molecules and so increasing the amount of enzymes at this point, has no affect. The graph plateaus off once again (THICK LINE ON GRAPH).

Catalase catalyses the breakdown of Hydrogen Peroxide into water and oxygen. You can then collect the oxygen produced per minute to measure rate.

The enzyme amylase catalyses the breakdown of starch to maltose. Its easy to detect starch using a solution of potassium iodide and iodine. You then time how long it takes for the starch to disappear by regularly sampling the starch solution and use the times to compare rates between different tests.

Describe method and the apparatus you'd use.

Mention what you're measuring (Dependent variable).

--> eg. the volume of gas produced per minute

Describe how you'd vary the Independent variable.

--> eg. possibly five different concentrations of enzyme

Describe the variables you are keeping constant.

--> temperature, pH, volume of solution etc.

Say that you need to repeat the experiment (twice or more) to increase reliability.

Mention you need a control.

--> eg. test tube containing the substrate but no enzyme.

An inhibitor is a substance that can slow down or stop a reaction. The inhibitor interferes with the reaction by binding to the enzyme thereby stopping the substrate binding instead. The two types of inhibitors are reversible and irreversible;

Irreversible - Heavy metal ions eg. Mercury (Hg+) or Silver (Ag+) are irreversible inhibitors that cause disulphide bonds within the protein structure to break. This leads to a permanent loss of structure and hence, activity. 

Other irreversible inhibitors such as cyanide, organophosphates or nerve gas permanently bind to the enzyme molecule which prevents the substrate binding. 

If the inhibitor has strong, covalent bonds, the inhibitor cannot be removed. It is irreversible.

Reversible - If the inhibitor has weaker hydrogen bonds or weak ionic bonds, then it can be removed. It is reversible. Reversible inhibitors can be split up into two types. Competitive and Non-competitive inhibitors.

Competitive inhibitors have a similar structure to the substrate molecule that binds to the active site of an enzyme.

They compete with the substrate for the chance to bind with the active site, but no reaction actually occurs.

They effectively block the active site so that not substrate can currently fit in.

The amount of product formed by the enzyme will eventually be the same; it just takes longer as the rate of reaction has been decreased. 

The effectiveness of an inhibitor depends on the relative concentrations of the substrate and the inhibitor. If theres a high concentration of the inhibitor and low concentration of substrate, it is more likely to bind to more active sites and it will reduce the rate of reaction significantly.

If the concentration of substrate is high and the concentration of inhibitor low, then more substrate will bind with the active sites available and the inhibitor will have a less dramatic effect on the rate of reaction.

A non-competitive inhibitor does not bind to the active site of an enzyme, it binds at another point on the enzyme structure.

The binding of the inhibitor alters the shape of the active site and so prevents the binding of the substrate.

They do not compete with the substrate because they have a different shape and bind elsewhere on the enzyme.

Increasing the concentration of substrate will not overcome this type of inhibition, the enzyme will still be inhibited.

You need to know an example of a poison for F212. You only need to know one and be able to describe its action. Choose from list below;

CYANIDE - Irreversible inhibitor that attached strongly to the copper containing prosthetic group of cytochrome c oxidase,  an enzyme that catalyses aerobic respiration.  Cells that cannot respire therefore die. Without the antidote you can suffer from unconsciousness, coma and death.

MALONATE - Inhibits succinate dehydrogenase which catalyses respiration.

ARSENIC - Inhibits pyruvate dehydrogenase which also catalyses respiration.

ORGANOPHOSPHATES AND NERVE AGENTS - Many of them inhibit acetylcholinesterase which normally removes the neurotransmitter acetyl choline after a nerve impulse has been transmitted across a synapse or across a neuro-muscular junction. Thus organophosphates disrupt normal nervous control by causing continuous stimulation of the nerve or muscle beyond the synapse.

You need to know an example of a medicine for F212. You only need to know one and be able to describe its action. Choose from list below;

PENICILLIN - Used to fight bacterial infections. Binds strongly to the active site of an enzyme (transpeptidase) that normally links peptidoglycan molecules together in bacterial cell walls. Consequently the bacteria cell bursts when it absorbs water.

SULPHONAMIDE - Bacteria use para-amino benzoic acid (PABA) to synthesise folic acid which is essential for growth. Sulphonamide drugs have a similar structure to PABA and so block the active site of the enzyme (dihydropteroate synthetase) needed to make folic acid.

RITANOVIR - An antiretroviral drug used to treat HIV infection. It binds to the active site and inhibits HIV protease, an enzyme used in the final assembly of new viral particles.

Others include ETHANOL and ANTABUSE (Disulfiram is the active ingredient).

Some enzymes need the presence of another molecule if they are to work. These molecules are known as cofactors.

Cofactors are non-protein molecules.

They modify the chemical structure of the enzyme in some way so that it can function more effectively.

There are three types of cofactors;

PROSTHETIC GROUPS

COENZYMES

ACTIVATORS

PROSTHETICS - Are organic molecules that form a permanent attachment to the enzyme eg. Haemoglobin contains the prosthetic group haem which contain iron and bonds permanently to the protein molecule. Haem enables haemoglobin to carry oxygen. It is also present as the prosthetic group in the enzyme catalase.

COENZYMES - Are small non-protein organic molecule. Unlike prosthetic groups, they do not bond permanently. Coenzymes help enzymes and substrates to bond with each other. The enzyme can only function if the coenzyme is present. Many coenzymes are derived from vitamins eg. NAD from the vitamin nicotinic acid. NAD is the coenzyme for a number of dehydrogenase enzymes. It acts as a hydrogen acceptor.

ACTIVATORS - Inorganic metal ions. They form a temporary attachment to the enzyme and change its active site so that the reaction is more likely to take place eg. the synthesis of any protein in our bodies requires magnesium as a cofactor. The reaction does not take place without it. eg.2 Manganese ions are cofactors found in hydrolase enzymes (enzymes that catalyse the hydrolysis of chemical bonds). Most metal activators are obtained from our diet.