An investigation into how concentration of enzyme affects rate of reaction

Christopher Kurwie - - Biology Sc1

11PS 26/04/2007

Sc1 – An investigation into how concentration of enzyme affects rate of reaction

AIM

The aim of this investigation is to see how different levels of enzyme concentration affect the rate of an enzyme controlled reaction. In this case, the substrate is pectin, found in the cell walls of apples, and the enzyme is pectinase.

VARIABLES

The variables in this experiment are:

temperature
pH
concentration of enzyme
concentration of substrate
time
inhibitors

The chosen variable is concentration of enzyme.

HYPOTHESIS

I predict that as the concentration of enzyme increases, the rate of the reaction will also increase. This will be in a directly proportional ratio, until a peak rate of reaction occurs where the rate cannot increase any further. This will produce a straight-line graph through the origin, which will eventually flatten off at peak rate. This is what I estimate the graph to look like:

Transmission

of light (%)

Enzyme conc.

This hypothesis can be summed up in one simple quotation from G.Toole and S.Toole, Understanding Biology:

“The rate of a reaction is directly proportional to the enzyme concentration. The addition of further enzyme cannot increase the rate of reaction and the

graph will tail off.”

My preliminary work showed this. Different concentrations of amylase where placed onto a petri dish of starch. The highest concentration of starch broke down the most starch in the allotted time.

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Transmission

of light (%)

Enzyme conc.

This hypothesis can be summed up in one simple quotation from G.Toole and S.Toole, Understanding Biology:

“The rate of a reaction is directly proportional to the enzyme concentration. The addition of further enzyme cannot increase the rate of reaction and the

graph will tail off.”

My preliminary work showed this. Different concentrations of amylase where placed onto a petri dish of starch. The highest concentration of starch broke down the most starch in the allotted time.

When an enzyme meets the substrate, the two combine. The enzyme reacts with its substrate, splitting it into two. It is then that it has been broken down. The enzyme is then free to combine with another substrate molecule and restart. The location where this occurs is called the ‘active site’. It stands to reason that the more enzyme molecules there are, the quicker this reaction will take place. If there is double the amount of enzyme molecules, the reaction will occur at double the speed. There will be however a maximum rate when the reaction cannot occur any quicker.

APPARATUS

The apparatus that I shall use is:

six cuvettes
colorimeter
apple juice (cloudy)
pectinase
water
syringe

METHOD

To measure the rate of the reaction, I plan to take readings of the amount of light that is transmitted thorough the cuvettes. As the enzyme breaks down the apple juice, it becomes clearer, and so more light is passed through. It is for this reason that I shall use a colorimeter to measure the transmission of light and hence the rate of the reaction.

I will take the apple juice and pectinase and place them in the cuvettes in the correct proportions. I wish to create 5 different concentrations, and a control. These are 0%, 20%, 40%, 60%, 80% and 100%.

I will set up 6 cuvettes each with 0.5ml of apple juice in them.
To these I will add the water and pectinase in the concentrations stated above.
After these are labelled, I will place each of these in the colorimeter, and an initial reading of the transmission of light will be taken. Between these, a colorimeter of water must be used to re-zero the apparatus.
Repeat this step after 30 minutes and measure the new levels of transmission of light.
I will then repeat the entire experiment three times, to try and remove any anomalous results that may occur.
To check that it is the enzyme that is causing this reaction, and not the water or any other factors, I will perform a control. This is a cuvette full of only water and apple juice, but no pectinase. If this changes in any way, I can be sure that my experiment is crucially flawed.

CONTROL OF OTHER VARIABLES

I will not alter the temperature, and I deem that any changes that occur will only be very minor.
The pH will be kept the same by using the same source of tap water and apple juice.
I cannot measure the substrate concentration, but I assume this to be the same throughout.
There will be no inhibitors in the experiment, unless they are in the water, I which case it is the same for all of the tests.

If all these are adhered to, then I can be sure that the experiment will be a fair one, and that all results gained will be just.

SAFETY

There are no dangerous chemicals I this experiment, but I plan to wear an apron at all times, as well as goggles in case of splashes. This should mean that it will be carried out safely.