Chapter 4

Enzymes are important because the processes necessary for life involve chemical reactions that need to happen very fast but not at high temperatures, so enzymes are used as biological catalysts which are globular proteins that interact with substrate molecules causing them to react much faster without the need for harsh conditions. Anabolic enzymes are those that synthesis large large polymer based components , these are synthesised and assembled in cells which form tissues, organs and organ systems. Catabolic enzymes are those that break down large organic molecules in order to release enegry needed for metabolic function, such as the dugestion of starch into glucose. 

Molecules in a solution collide randomly, for a reaction to occur the molecules need to collide at the right orientation along with having enough energy for the reaction to occur. enzymes help molecules collide so reduce thre activation energy. Each enzyem catalyses one biochemical reaction.

Lock and key model: An area within the tertiary structure of the enzyme has a shape complementary to the shape of a specfic substrate molecule - active site. Only the correct substrate will fit into the active site. When a substrate is bound to the active site an enzyme-substrate complex is formed, the substrate(s) then react and the product(s) are formed in an enzyme-product complex. The product(s) are released, leaving the enzyme unchanged for more reactions. The substrate is held in such a way that the right atom groups are close enough to react, the r-groups within the active site will also interact with the substrate, putting strain on the bonds in the substrate and so aiding the reaction.

Induced fit hypothesis: New research suggests that the active site changes shape slightly as the substrate enters, the initial interaction between them is weak but they rapidly induce changes in the enzymes tertiary structure that strengthen the binding and put strain on the substrate molecule and weakens bonds in it, so lowers the activation energy for that reaction.

Enzymes such as those involved in the synthesis of polymers work inside the cells so are intracellular. For example hydrogen peroxide is a toxic by-product of many metabolic pathways but is broken down to oxygen and water quickly to prevent its accumulation by the enzyme catalase.

All of the reactions inside cells need substrates which need to be constantly supplied, nutrients present in diet or enviroment supply these materials. They often come in polymers that cannot enter the cells directly so need to be broken down into smaller components, enzymes are released from cells to break down these nutrient molecules into smaller ones in the process of digestion, these are extracellular enzymes. Single celled organisms release extracellular enzymes into their immediate enviroment to break down larger molecules, multicellular organisms digest nutrients and absorb the smaller components into the bloodstream where they will be transported to the cells for reactions, for example amylase breaks down starch into maltose in the saliva, and trypsin hydrolyses peptide bonds to break dwon proteins in the small intestine.

Digestion of starch: It begins in the mouth and continues in the small intestine, it is digested in 2 steps involving 2 different enzymes.

1. Starch polymers are broken down into the disaccaride maltose by amylase that is secreted from the salivary gland and in thr pancreatic juice released into the small intestine.

2. Maltose is then broken down into glucose by maltase in the small intestine.

Digestion of proteins:

Trypsin is is a protease which is a type of enzyme that catalyses the digestion of proteins into smaller peptides, which can then be further broken down into smaller peptides by other proteases. Trypsin is produced in the pancreas and released into the pancreatic juice where it acts on the proteins in the small intestine. The amino acids produced are absorbed by the cell lining of the digestive system and then absorbed into the bloodstream. 

Increasing the temperature of a reaction increases the kinetic energy the particles have and so will lead to more successful collisions in an enzyme controlled reaction. The temperature coefficient is a measure of how much the rate increases with a 10°C  inccrease in temperature, for most enzyme controlled reactions Q10 wil be 2 meaning the rate doubles with an increase of   10°C.

Denaturation: As the temperature increases it increases the vibrations of the protein bonds and structure causing them to strain and then break, causing change in the precise tertiary structure of of the protein - it is said to be denatured. When an enzyme is denatured the active site changes shape and is no longer complementary to the substrate and so will no longer function as a catalyst.

Optimum temperature: This is the temperature that has the highest rate of activity, it can vary significantly depending on the enviroment the enzyme is present in. Once an enzyme has been denatured the decrease in rate is rapid - there only needs to be a tiny change in structure will make it no longer complementary.

Temperature extremes: Some organisms can survive harsher temperatures, thermophiles have enzymes with far more bonds to make them more resistant, psychophiles have fewer bonds so their active sites are more flexible.

Enzymes are also affected by change in pH, which refers to a change in hydrogen ion concentration and alter the ionic, hydrogen and polar bonds in the tertiary structure in proteins. The active site will only be the right shape at a particular hydrogen ion concentration - optimum pH. When pH changes the structure is altered, but if it returns to the optimum then the protein will resume its function as a catalyst - renaturation. When the pH changes more significantly the structrue is irreversibly changed. Hydrogen ions interact with polar and charged r-groups, so a change in pH also affects the interaction of r-groups with each other. The more H ions present (lower pH) the less r-groups are able to interact with each other leading to bonds breaking and the shape changing and vice versa for a higher pH. Enzymes only function in a very narrow pH range.  

When the concentration of a substrate is increased the number of substrate molecules increases leading to a higher collision rate, and this is the same when you increase the enzyme concentration as the number of enzyme-substrate conplexes will increase, it wil only increase up to a certain point (V max). At this point all of the active sites are occupied so no more complexes will be formed until products are released from the active sites, or the amou nt of substrate will become the limiting factor if the enzyme concentration is increased. 

A serial dilution is a stepwise dilution of a stock solution of a known concentration to produce a range of concentrations, they are used to investigate the concentration of an enzyme of substrate in a reaction. For example adding 1 ml of stock solution to 9 ml of distilled water gives a 1/10 dilution. To investigate the concentration of catalase you can grind up potato to mkae a solution and then carry out a serial dilution to show relative concentrations of the enzyme, the effect of these could be d=seen by adding the same amount of hydrogen peroxide to each one.

Different steps of reaction pathways are controlled by different enzymes, controlling the activity of enzymes at crucial points in these pathways regulates the amount of product formed, enzymes can be activated by cofactors, or inactivated by inhibitors, which are molecules that orevent enzymes from carrying out their normal function of catalysis or slow them down, there are two types of inhibition, competitive and non competitive.

• A molecule or part of a molecule that has a similar shape to the substrate of an enzyme can fit into the active site.
• This blocks the substrate from entering the active site, preventing the enzyme from catalysing the reaction - cannot carry out its function so is said to be inhibited.
• The non-substrate ,olecule that binds to the active site will compete with substrate molecules to bind to the active sites, which will reduce the amount of product being made, and the degre of inhibition will depend on the relative concentrations of substrate, inhibitor and enzyme.

Competitive inhibitors reduce the rate of a reaction but they do not change the V max of the enzyme it inhibits. Statins are an example of competitive inhibitors that inhibit synthesis of cholesterol. Aspirin irreversibly inhibits the active sites of COX preventing the synthesis of thromobxane which is the chemical responsible for producing pain.

• The inhibitor binds to the enzyme at a location other than the active site - called an allosteric site.
• The binding of the inhibitor causes the tertiarty structure of the enzyme to change meaning the shape of the active site is also changed.
• This results in the active site no longer being complementary to the shape of the substrate molecule so it is unable to bind to the enzyme.
• The enzyme cannot carry out its function so it is said to be inhibited.

Increasing the concentration of an enzyme or substrate will not overcome the effects of a non competitive inhibitor, but increasing its concentration will decrease the rate of reaction further as more active sites become available. \Example include insecticides which are permament inhibitors and cause paralysis and death when ingested. Proton pumps are used to treat long term indigestion by irreversibly blocking an enzyme system responsible for sectreting hydrogen ions into into the stomach. 

End product inhibition is the term used when the product of a reaction acts as an inhibitor to the enzyme that produces it. This serves as a negative feedback mechanism that prevents excess product from being made and resources are not wasted. The first step of respiration involves the addition of two phosphate groups and is catalysed by the enzyme PFK, this enzyme is competitively inhibited by ATP, and therefore ATP regulates its own production. When levels of ATP are high more of it will bind to the allosteric site on PFK and so glucose wont be broken down and the reaction will not occur. As ATP is used up, less binds to PFK and the enzyme is able to catalyse the addition of the second phosphate group leading to the production of more ATP.

Some enzymes need a non protein helper component in order to carry out their function, they may transfer atoms of groups or form part of the active site, these components are called cofactors, or if it is organic then it is a coenzyme. Inorganic cofactors are obtained via the diet as minerals, eg amylase has a chlroide ion in its active site. Many coenzymes are derived from vitamins, eg B3 is used to synthesise NAD which a coenzyme responsible for transfer of hydrogen atoms between molecules involved in respiration. 

Prosthetic groups are a type of cofactor, whilst some cofactors are temporarily bound to the enzyme to activate them, prosthetic groups are tightly bound and form a permanent part of the protein's structure. Eg zinc ions form an important part in the enzyme carbonic anhydrase, an enzyme necessary for metabolism of carbon dioxide.

Many enzymes are produced in an inactive form, known as precursor enzymes, particularily those that can cause damage within the cells producing them or to the tissues when they are released. Precursor enzymes often need to undergo a change in tertiary structure, specifically to the active site to be activated, this can be achieved by the addition of a cofactor. Before it is added the enzyme is called a apoenzyme, when it is added it is called a holoenzyme. Sometimes that change in tertiary structure is brought about by the action of another enzyme, such as a protease which cleaves bonds in a molecule. A change in pH or temperature can result in a change in tertiary structure to activate a precursor enzyme - zymogens and poenzymes. For example when inactive pepsinogen is released into the stomach to digest proteins, the acidic pH transforms it into the active enzyme pepsin, without it damaging the other body cells.