Dear readers, this is a concluding post to my previous post on ENZYMES AND METABOLISM.
I will start my discussion today on types of enzymes. Although there are many different enzymes, each of them can be put into one of six main categories according to the type of reaction they catalyse:
- Oxidoreductases: These catalyse oxidation and reduction (redox) reaction. In aerobic respiration, most of the cell's ATP is generated by redox reactions.
- Transferases: These catalyse the transfer of a chemical group from one compound to another, for example, the transfer of an amino group from an amino acid to another organic acid in the process of transamination.
- Hydrolases: These catalyse hydrolysis (splitting by use of water) reactions. Most digestive enzymes are hydrolases.
- Lyases: These catalyse the breakdown of molecules by reactions that do not involved hydrolysis.
- Isomerases: These catalyse the transformation of one isomer into another. For example, the conversion of glucose 1,6-diphosphate into fructose 1,6-diphosphate. This is one of the first reactions in glycolysis, the first stage of respiration.
- Ligases: These catalyse the formation of bonds between compounds often using the free energy made available from ATP hydrolysis. DNA ligase, for example, is involved in the synthesis of DNA.
FACTORS THAT AFFECT ENZYME ACTIVITY
Enzymes are proteins and their function is therefore affected by: temperature, pH. substrate concentration, enzyme concentration and inhibitors.
For a non-enzymic chemical reaction, the general rule is the higher the temperature, the faster the reaction. This same rule holds true for a reaction catalysed by an enzyme, but often only up to about 40 to 45°C. Above this temperature, enzyme molecules begin to vibrate so violently that the delicate bonds that maintain tertiary and quaternary structure are broken, irreversibly changing the shape of the molecule. When this happens the enzyme can no longer function and we say it is denatured.
The effect of temperature on a reaction can be expressed by the temperature coefficient, commonly known as the Q10, where t is the chosen temperature, the formula for the Q10is:
Rate of reaction at t + 10°C ÷ Rate of reaction at t°C
In an illustration to calculate Q10. To avoid denaturing the enzyme, the values for living organisms need to fall in the range 4 to 40°C. So we have chosen t as 20°C.
Q10 = rate at 30°C ÷ rate at 20 °C = 4/2 = 2
In practice, most enzymes have a Q10 between 2 and 3. A value of 2 means that the rate of reaction doubles with a 10°C temperature rise, 3 means that it triples. Some organisms have enzymes that are less sensitive to heat than those found in mammals. For example, certain bacteria can survive in hot volcanic springs and deep sea hydrothermal vents at temperatures of over 90°C. So their enzymes must be active at these extreme temperatures.
Stable enzymes for washing powders
Enzymes are unstable, particularly at high temperature, so their commercial usefulness is limited. Many industrial processes need to take place in ‘unnatural’ environments, at high temperature and extremes of pH. But there are organisms that can thrive at high temperatures. Heat-loving bacteria have thermostable enzymes that are also more resistant to extremes of pH and other unfavourable conditions, such as organic solvents.
Genes that code for some thermostable enzymes have been transferred into bacteria that are easy to grow in large quantities, using recombinant DNA technology. As the bacteria that have received a particular gene multiply, the gene is translated into protein and large amounts of the enzyme are produced for commercial use.
An example of a thermostable enzyme is the alkaline protease subtilisin, the famous stain digester in biological washing powders. This ernzyme is produced on a grand scale by the bacterium Bacillus subtilis, and is active in alkaline environments, and so is compatible with the other ingredients in washing powder. It is active at temperatures up to 60°C, allowing it to be used in a wide variety of wash programmes.
Like other proteins, enzymes are stable over a limited range of pH. Outside this range, enzymes are denatured. Free hydrogen ions (H+) or hydroxyl ions (OH-) affect the charges on amino acid residues, distorting the three-dimensional shape and causing an irreversible change in the protein’s tertiary structure.
Enzymes are particularly sensitive to changes in pH because of the great sensitivity of their active site. Even if a slight change in pH is not enough to denature the molecule, it may upset the delicate chemical arrangement at the active site and so stop the enzyme working.
Most enzymes are intracellular: they work inside cells, and their optimum pH is around 7.3 to 7.4. Most organisms have buffer systems that resist changes in pH, and many are able to excrete excess acid or alkali. In a figure showing some enzymes that have optimums of pH that are distinctly acid or alkaline. These include digestive enzymes that normally work in the stomach or inside lysosomes. We describe enzymes that work outside cells as extracellular.
The rate of an enzyme-controlled reaction increases as the substrate concentration increases, until the enzyme is working at full capacity. At this point, the enzyme molecules reach their turnover number and, assuming that all other conditions such as temperature are ideal, the only way to increase the speed of the reaction even more is to add more enzyme.
In any reaction catalysed by an enzyme, the number of enzyme molecules present is much smaller than the number of substrate molecules. Looking at the table below, which shows the turnover numbers of some enzymes, and you can see that one molecule of an enzyme can convert millions of substrate molecules into products every minute.
When there is an abundant supply of substrate, the rate of reaction is limited by the number of enzyme molecules. In this situation, increasing the enzyme concentration increases the rate of reaction.
A table showing the turnover number of some enzyme
|Carbonic anhydrase||36 000 000|
|Catalase||5 600 000|
Inhibitors slow down or stop enzyme action. Usually, enzyme inhibition is a natural process, a means of switching enzymes on or off when necessary. Inhibition tends to be reversible, and the enzyme returns to full activity once the inhibitor is removed. Many ‘external’ chemicals, such as drugs and poisons, can inhibit particular enzymes. This type of inhibition is often non-reversible. Reversible inhibitors are either competitive or non-competitive.
Competitive inhibitors compete with normal substrate molecules to occupy the active site. The inhibitor molecules must be a similar size and shape to the substrate to fit the active site, but cannot be converted into the correct product. Effectively, competitive inhibitors ‘get in the way and reduce the number of interactions that can happen between enzyme and substrate.
Non-competitive inhibitors bind to the enzyme away from the active site but change the overall shape of the molecule, modifying the active site so that it can no longer turn substrate molecules into product. Non-competitive inhibition has this name because there is no competition in the active site. The presence of a non-competitive inhibitor has the same effect as lowering enzyme concentration: all inhibited molecules are taken out of action completely.
Irreversible inhibitors bind permanently to the enzyme, rendering it useless. For obvious reasons organisms rarely produce this type of inhibitor for their own enzymes. However, irreversible inhibitors are splendid weapons to use against other organisms. A wide variety of natural toxins are irreversible inhibitors, as are many pesticides. Cyanide is an irreversible inhibitor of cytochrome oxidase, one of the enzymes involved in respiration. Organisms poisoned with cyanide die because they are deprived of ATP, their immediate energy source.
How enzymes help babies digest cow’s milk
When babies are first born, their intestines don’t behave at all like those of an adult. So that they can absorb and make use of important antibodies in their mother’s breast milk, they have intestinal walls with a structure that resembles a string vest. It has lots of large pores through which whole proteins can pass, without being digested into their component amino acids.
Everything is fine, as long as the baby is fed breast milk, but some mothers cannot breast-feed for very good reasons. What happens then? A hundred years ago, babies that couldn’t feed from their mother usually fell behind in their growth, sickened and often died. Cow’s milk does not contain the same proteins as human milk, and it also has a completely different balance of minerals such as sodium. Feeding tiny babies on untreated cow’s milk leads to a generalized immune response against the cow proteins and can also damage the delicate kidneys, which are not able to cope with large amounts of sodium.
Today, many babies are fed on infant formula milk and they thrive just as well as babies that are breast-fed (although the natural method is still considered the best). Enzyme technology has had a lot to do with this – enzymes have been used in the production of infant formula milk from cow’s milk for over 50 years. Manufacturers used proteases to digest the milk proteins into short peptides and individual amino acids, so preventing whole cow proteins from entering the baby’s system. The ‘pre-digested’ cow’s milk is also adjusted for fat and mineral content, and today’s formula milks are as close to human milk as possible. Whether biotechnology will ever manage to add all the antibodies, cells, human enzymes and other components of naturally produced human milk remains a question for future.
How inhibitors help to control metabolism
Many metabolic pathways are self-controlling: when a substance is needed, a particular pathway is activated to produce it. When enough has been produced, the pathway is deactivated. This happens because some enzymes in a metabolic pathway are inhibited by the end product. If too much product begins to accumulate, this inhibits one of the enzymes in the pathway. When the product is once more in short supply, the inhibition is lifted and the pathway becomes active again. This self-regulation is an example of negative feedback. This is a fundamental principle, important in homeostasis.
After reading through all my series of posts written on Enzyme starting from this and this concluding post, you should know and understand the following:
Enzymes are globular proteins with a precise, but delicate, three-dimensional shape maintained by ionic forces and hydrogen bonds. During a reaction, the substrate fits into a region on the enzyme surface called the active site. Enzymes are specific: each enzyme catalyses one particular reaction. Enzymes speed up reactions by lowering the activation energy needed to get the reaction started.
Between 4°C and 40°C. The rate of an enzyme-controlled reaction increases between two-and threefold for every 10°C rise in temperature. Increase in temperature beyond 40 °C usually denatures the enzyme. Activity is lost when its three-dimensional structure is destroyed.
Most enzymes have an optimum pH. For intracellular enzymes this is usually about 7.35. Extracellular enzymes, such as those found in digestive juices, may have optimums of extreme pH values. The rate of an enzyme-controlled reaction is limited by the supply of substrate, enzyme or the enzyme cofactor. Inhibitors are substances that slow down or stop enzyme activity. They may be reversible or non-reversible. Reversible inhibitors may be competitive or non-competitive.
Competitive inhibitors tend to be similar in structure to the substrate and compete for the active site. Non-competitive inhibitors do not bind to the active site itself but alter the shape of the enzyme so that the active site is no longer functional.
Thank you all for coming.