Enzymes

Enzymes as catalysts lowering activation energy through the formation of enzyme-substrate complexes.

The important properties of enzymes are summarised in this section.

Enzymes are three-dimensional biological catalysts, globular protein molecules that are only produced in living organisms.

Enzymes possess a small region (typically only 20 amino acids or so in size) called the active site where the reaction occurs.

Enzymes are specific in the reactions that they catalyse, that is, one enzyme-one substrate. An enzyme may, however, act on closely related substrate molecules (often with different efficiencies). One type of molecule may also be the substrate for several different enzymes.

Enzyme activity conforms to the induced fit model. The substrate molecule induces a slight change in the shape of the active site to allow the substrate molecule to fit perfectly. The change in shape of the active site facilitates the reaction. After the reaction is complete, the product is released, the active site resumes its normal shape and the enzyme is free to attach to more substrate.

Enzymes are proteins, and are therefore affected by the properties of proteins. This means that their activity and/or structure will be affected by changes in conditions such as temperature and pH.

The rate of an enzyme-catalysed reaction is affected by:

• the concentration of the enzyme;
• the concentration of the substrate.

Since enzymes are catalysts, they:

• are required only in relatively small amounts;
• remain unchanged at the end of a reaction.

The properties of enzymes relating to their tertiary structure.

Remember that the protein is actually a really long chain of amino acids, strung together and forming a structure which depends upon the bonds (especially hydrogen bonds) between atoms in the molecule. Because this produces a globular protein with a tertiary structure, and this structure depends upon the order of the amino acids, then a specific shape is formed by the protein. This protein will be different from other proteins because other proteins have a different order in their amino acids, and therefore a different shape will be formed.

One specific part of the enzyme will have a shape which is complementary to the substrate. This means that, in the same way that a glove is shaped to fit a hand, so the active site is shaped to fit the substrate. If there is a mutation in the DNA so that one of the bases is changed, then a different amino acid may appear in the protein. If this happens to be one of the amino acids in the section of the protein which forms the active site, then the active site may change shape, so it no longer fits the substrate. This in turn means that there may be no enzyme for a particular reaction in the body, and that can be catastrophic. Mutations in genes coding for enzymes are a cause of many diseases.

If you want to go into a bit more detail, then you might like to know that the active site isn't actually complimentary to the shape of the subtrate, as such. We actually look at the process as a whole. Generally speaking we can say that a substrate has to undergo a reaction to form a product, and on the way it will form some kind of intermediate. The image on the right represents this as a circle turning into a star, and a cross between the two in the middle.

An enzyme works because the active site is actually complementary to the intermediate of the reaction. The substrate basically forms the intermediate and therefore is able to fit into the active site of the enzyme. The fact that the active site is complimentary to the intermediate 'encourages' the substrate to go through the reaction forming the intermediate. However, as it carries on the reaction to form the product, the shape of the product is no longer complimentary to the active site, and therefore the product is forced out of the active site, leaving both the product and the enzyme to do their own business - and allowing the enzyme to remain unchanged.

It's a very clever business, but it's also very complicated. All you really need to understand is that the active site is complimentary to the substrate, but understanding its relation to the intermediate helps to understand how the enyme works.

Importantly, like evolution, this is all theory. None of it has actually been proven because it's difficult for us actually to know. Unlike evolution, the lock and key theory is widely accepted because it combines plausibility with scientific evidence, but it's difficult to say for sure exactly how much enzyme function relates to complementary substrates...

Description and explanation of the effects of temperature:

Temperature is one of those marvellous things - once you understand that heat is simply more energy in the particles, you've understood the most complicated part. Well, almost anyway. The thing is, it doesn't really get that more complicated. The principles of temperature are pretty much the same for everything. So, when it says in the rates of reactions section that temperature increases reaction rate, you would be right in thinking that temperature also increases the activity of an enzyme.

It all relates to the movement of the particles again. You have your substrate whizzing about (lets assume everything is dissolve in solution) and you've also got your enzyme there as well. Well, ideally you'd like the enzyme molecules and substrate molecules to meet as many times as possible, because the more times they hit each other, the more likely it is that the substrate will slot into the active site, leading to the product being formed.

The higher the temperature, the more energy the molecules have, so they'll be moving more quickly, and are more likely to collide successfully,

BUT

the problem you get is that an enzyme is a biological molecule, and in the same way that we don't like temperatures to get too hot, enzymes don't respond well to high temperatures. When the temperature rises too much, proteins get denatured, which means that bonds between the functional groups of amino acids break, and the protein changes shape. This means that the enzyme's function, which relies so much on its shape, is lost, and the enzymes become useless. So, at a certain temperature, the enzymes stop working, and any further increase in temperature will not help the enzymes activity any more. In the body, this is often around 37ºC.

So as you increase temperature, the activity of an enzyme will increase, until you reach the optimum value. After that, any further increase in temperature will result in denaturation of the enzyme, and a steep drop in activity.

Description and explanation of the effects of competitive inhibitors:

Competitive Inhibition is where the inhibitor is a molecule which has a similar shape to the molecule which is supposed to be binding to the active site. In the case of enzymes, a competitive inhibitor may have the same shape as that of the substrate, but it doesn't react in the same way. Rather than turning into the product, it simply uses up time and prevents the substrate from getting to the active site. It blocks the way.

Description and explanation of the effects of non-competitive inhibitors:

Non-competitive Inhibition exists when a molecule binds to a different site on the protein, rendering it inactive. Sometimes it does this before the substrate reaches the active site, sometimes afterwards, but in either case it stops the protein doing its job, and prevents a product being formed.

In both competitive and non-competitive inhibition, it is possible to have both reversible and irreversible inhibitors. As the name suggests, a reversible inhibitor does not have a permanent effect - it will stop the protein doing what it is supposed to do, but it will move off again and allow the protein to function later on; an irreversible inhibitor, on the other hand, permanently renders the protein inactive, so it will have to be replaced by a brand new one - the inhibitor will not budge.

Inhibitors in general

If there is an inhibitor about, then changing the concentration of substrate won't change the rate of the reaction in the same way as before. The problem you have is that something is stopping the enzyme doing its job properly, and so although increasing the substrate concentration does increase the reaction rate, it simply cannot do so to the same extent.

The way in which the situation differs depends upon which kind of inhibitor you're using. If the inhibitor is non-competitive (that is, it's acting on a site other than the active site, then the rise in reaction rate in comparison to substrate concentration will be the same shape but squashed. Due to the inhibition, the enzyme simply cannot achieve the same level of activity; the optimum substrate concentration will still be the same because it will require the same concentration of substrate to fill all of the active sites on the enzymes, but it won't produce the same level of activity because it's all being inhibited.

A competitive inhibitor is slightly different because this time you do have some hope of reaching the normal level of enzyme activity. At low substrate concentrations, there will be a comparitively large proportion of inhibitor molecules floating about - that is, you may have 2 substrate molecules and 4 inhibitor molecules for 5 enzymes. Obviously the inhibitor molecules are more likely to get into the active sites. However, if you increase the concentration of substrate molecules, you may have 7 substrate molecules and 4 inhibitor molecules for 5 enzymes. This time the substrate molecules are more likely to get into the active sites - but you'll still have some inhibition.

Eventually you'll have so many substrate molecules that the inhibitor molecules are much, much less likely to get into the active sites, and so the substrate molecules will fill the active sites. This is effectively the same situation you had when the optimum substrate concentration was achieved before - all the active sites are filled with substrate, so the enzyme is doing the best job it can.

The difference with a competitive inhibitor in comparison to a non-competitive inhibitor is that, if all the active sites are filled with substrate, there's no inhibition going on, and therefore the maximum reaction rate will be achieved. So, it is possible to overcome the effect of a competitive inhibitor, but you have to get a much higher substrate concentration than usual.

Description and explanation of the effects of pH:

There is an optimum value for lots of things. There is an optimum temperature for enzyme activity - there is an optimum speed for efficient fuel consumption in a car. In the same way, there is an optimum pH for each and every enzyme. At a pH above this optimum, the enzyme's activity will be reduced and therefore the reaction rate will be lowered; at a pH below this optimum, the enzyme's activity again will be reduced and lower reaction rates result.

The obvious question is, why? Why does changing the pH - changing the concentration of hydrogen ions in the solution - have such an effect on the enzyme?

To begin with we have to rethink the idea we have of the active site. It's not simply a case of the molecule forming a particular shape. Unfortunately it's a bit more complicated than that. When a substrate molecule slots into the active site of an enzyme, it doesn't just fit snuggly like a glove - although that is a reasonable simplification of it. Instead, the active site is made up of a series of groups of atoms which form temporary bonds to the substrate. These groups are the functional groups of the amino acids in the protein chain, and they have an important effect on the shape of the enzyme. If they're in a different order, the substrate won't temporarily bond to the enzyme, but the shape will also be different, so it's often easier just to think about it from the point of view of shape.

However, if we want to understand the effect of pH, we really need to be able to understand the way in which is it the position of the atoms which has such an important effect. This means that if the group changed - even if the shape stayed the same - the substrate may not be able to bond to the active site. This is what happens when the pH changes. If the pH increases, this means there are fewer hydrogen ions in solution. This can lead to reactions taking place that alter the functional groups of the amino acids, which in turn leads to the enzyme changing shape. Similarly if the pH decreases, there are more hydrogen ions in solution, leading to the possibility of hydrogen ions causing other group changes.

If the functional groups change, the shape will change, and the active site will no longer fit the substrate. This is what happens when pH changes. If the pH is changed sufficiently, the enzyme will be completely altered due to this effect, and it is said to be denatured. However, unlike the effect of extreme heat, which causes the enzyme to be irreparably damaged, denaturation due to pH change is reversible. Restore the pH to its original level, and the enzyme will return to its original capability.

Description and explanation of the effects of substrate concentration:

If you want an enzyme to work best, then you need it to have as fast a turnover as possible. You need as many substrate molecules to meet up with that enzyme within a given time as possible. So, it makes sense that if there are more of the substrate molecules in a given space, the enzyme can meet up with more of them in a given time!

This is effectively what is happening when you alter the substrate concentration . The enzyme, busily going about its catalytic work, will simply hang around until a substrate molecule collides with it appropriately. In the same way that increasing the temperature increases the chances of a favourable collision, so increasing the substrate concentration increases the chances - because there are simply more substrate molecules kicking about!

Unfortunately this doesn't carry on forever. Eventually you reach an optimum value - if you increase the concentration of the substrate passed this point, it makes no difference.

Why?

It's all to do with the number of available sites. Yes, enzymes are kicking about, 'waiting' for substrate molecules to jump in so they can do their work. However, there's only so much work that each enzyme molecule can do. Eventually you'll reach a stage where there are more substrate molecules than there are enzyme molecules to work their magic - so, as the image hopefully shows, you've got too many substrate molecules for any extra to make any difference. If you raise the concentration any further, it will make no difference because there are no extra enzyme molecules to meet this supply.

This is, of course, assuming you keep everything the same except for the substrate concentration. So, increasing the substrate concentration increases the activity of the enzyme, up to a point, after which increasing the substrate concentration has no effect.

Enzymes - an interactive site

How enzymes work

The Biology Project - Energy, Enzymes and Catalysis