the effect of substrate concentration on the rate of an anzyme reaction

The effect of substrate concentration on the rate of an enzyme reaction

Pilot and main experiment

Aim:

This is an experiment to investigate how the concentration of the substrate Hydrogen Peroxide (H2O2) affects the rate of reaction of the enzyme Catalase, in yeast cells.

Background theory:

An Enzyme is a globular protein, which is capable of starting a chemical reaction, which involves the formation or breakage of chemical bonds. The feature of Enzymes are to speed up the rate of reaction, it lowers activation energy (the higher the activation energy, the slower the reaction will be) , denatures if the temperature is too high and is a biological catalyst. Enzymes increase the rate of biochemical reaction to more than a billion times their normal rate, without being changed or used in the reaction. This is why they can be reused and even small concentrations of enzymes can be very effective. Enzymes cannot make reactions occur which otherwise wouldn't happen, and they do not alter the final amount of product formed. In addition they are very specific and they won't catalyse just any old reaction, only those which are suited for the enzyme.

Enzymes formed and retained in the cell are known as intracellular enzymes, like Catalase, and may occur either in the cytoplasm or the nucleus of the cell. Certain other enzymes produced in the cell are packaged to be secreted from the cell, and work externally, including most digestive enzymes. These enzymes are called extracellular enzymes. Furthermore all enzymes contain an active site, a depression in the enzymes molecule, where a specific substrate molecule can fit into exactly and bind to. The active site is small and usually 2 to 20 amino acids. All enzymes have a substrate molecule, which is the same shape as its the active site. When substrate binds to the enzyme's active site, it forms an enzymes- substrate complex, which produces the product(s). Enzymes are very specific and this is because the enzyme and substrate shape fit exactly together, this is called the lock- and- key model which was suggested by Emil Fischer in 1894. Most enzymes, when the substrate slots into the active site, change their shape slightly, so that it can accommodate the substrate better- this is called induces fit. Enzymes only perform one type of reaction, and only have one specific type of substrate to do this. The enzyme Catalase catalyses the breakdown of Hydrogen Peroxide:

Hydrogen Peroxide ----------------> Water + Oxygen

2 H2O2 ----------------> 2H2O + O2

Catalase is an enzyme found in food such as potato and liver, and is used to remove Hydrogen Peroxide from cells, as it is the poisonous by-product of metabolism. There are two types of enzyme-substrate metabolic reactions: anabolic and catabolic. An anabolic reaction is when there are two-part substrate which is built together to form a new molecule. A catabolic reaction is where one substrate is broken down to produce two products. There are four factors that can affect the rate of an enzymes reaction: The pH, the temperature, the concentration of the enzyme solution and the concentration of the substrate solution. The pH can affect the rate of an enzyme-catalysed reaction by affecting the bonds in the secondary and tertiary structure of the enzyme's active site. If the bonds are broken, the shape of the active site can be distorted, so the substrate molecule cannot bind to the enzyme causing no enzyme-substrate complex, and therefore producing no products. Each enzyme has an optimum pH. Most enzymes work best in neutral or slightly alkaline conditions. The temperature affects the rate of reaction. If you increase the temperature you increase the amount of energy, therefore more enzyme and substrate molecules will collide more often, increasing random movement. The more collisions there are, the higher the chance is that a substrate molecule will fit an active site. The change in rate of reaction for each 10 C rise in temperature is called the temperature coefficient,Q10: Q10=( rate of reaction at X+ 10 C)/( rate of reaction at X C)

Each enzyme has an optimum temperature, the rate of reaction starts to decrease as the actie site can start to distort, causing no enzyme-substrate complexes and producing no product, as the substrate cannot bind. At very high temperatures the number of collisions may be very high but without the correct shape of the active sites, no products can be formed. At these very high temperatures the enzyme can become denaturated. This means the enzyme's active site has become permanently damaged. The concentration of the enzyme solution also affects the rate of an enzyme reaction. If there are more enzyme molecules than substrate molecules, then the substrate would be a limiting factor, as there would not be enough substrate to continue forming products. When all the active sites are in use, the optimum rate will have been achieved. But the reaction will take place very quickly and then finish if no more substrate is added because the substrate will run out. In the same way the concentration of the substrate affects the rate of reaction, only in this case the limiting factor is the lack of active sites. The rate of this reaction will be slow as there are not enough active sites. The optimum rate occurs when all of the active sites are in use.

Molecules have to collide with a certain minimum energy, Ea, for there to be a chance of reaction. This minimum energy is called activation energy and it varies from one reaction to another. Activation energy has units of kJmol .

Risk assessment:

Hydrogen Peroxide can be corrosive if strong, and irritant to the eyes and skin. For solutions less than 18 'vol' H2O2 there is a low hazard. For all solutions to or greater than 18 'vol' but less than 28 'vol' H2O2 the hazard is irritant to eyes and skin. Eye protection should be worn during the whole experiment and it should be handled with care. Contact with skin should be avoided and clothing should be protected. Any spills should be cleaned immediately. All Enzymes are potential allergies and can irritate membranes in the eye or nose if ...

the effect of substrate concentration on the rate of an anzyme reaction

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