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Aim: -
To investigate the effect of enzyme temperature on the activity of the enzyme Trypsin on the substrate Casein.
Introduction: -
Casein is an insoluble protein found in powdered milk, such as Marvel, which forms a white or cloudy suspension (not dissolved). A white suspension of Marvel milk in water clears if a protein-digesting enzyme is added. The enzyme used in this experiment is trypsin. ... This reaction is catalysed by proteolytic enzymes (proteinases) such as trypsin. Trypsin is released into the small intestine in alkaline conditions. ... Trypsin acts with other proteinases to break down proteins to peptides and amino acids. ... Trypsin is produced in an inactive form by the pancreas. ... Each is active only against the peptide bonds in protein molecules that have carboxyl groups donated by certain amino acids. For trypsin these amino acids are arginine and lysine. Trypsin is the most discriminating of all the proteolytic enzymes in terms of the restricted number of chemical bonds that it will attack. ... Trypsin is an enzyme that acts to degrade proteins. It breaks down the casein into polypeptides which are soluble. ...
Enzymes like trypsin are biological catalysts that speed up a reaction without being used up. ... One type of enzyme will react with only one type of substrate. The protein has an area on the surface where the reaction occurs called the active site. ... The active site of one type of enzyme has the same arrangement of amino acids; therefore it has a highly specific shape. Usually there is only one active site on each enzyme and only one type of substrate will fit into it.
Trypsin catalyses the hydrolysis of peptide bonds but due to its shape, the active site of trypsin only splits bonds after a basic or straight chain amino acid. ...
A number of factors affect the rate of activity of trypsin on casein:
pH
Temperature of enzyme and substrate
Concentration of enzyme and substrate
*surface area is usually a factor, however, in this investigation both the substrate and enzyme are liquids. ... This is because the heat provides the enzyme and substrate with kinetic energy, therefore producing more collisions. ... Above the optimum temperature, the enzyme is denatured as it is a protein and its structure begins to break down due to excessive heat. ... The active site will be disrupted if the tertiary structure breaks down and the enzyme will be denatured.
If the enzyme concentration is too low there is great competition for active sites and there are not enough to bind to, making the reaction rate extremely slow. If there are more enzymes, there are more active sites to bind to so that more enzyme-substrate complexes can be formed. Eventually increasing the enzyme concentration will have no effect as there is not enough substrate to bind to its active site, and the substrate concentration becomes a limiting factor. ... If more substrate is added then more enzyme-substrate complexes can be formed, speeding up the rate of reaction. However, too much substrate means there will be no effect as the active sites will be saturated and no more enzyme-substrate complexes can form. ...
Dependant: activity of the enzyme, which will be measured using a stopwatch. ... It is inversely related, as the more time it takes to clear, the less enzyme activity there is. I will carry out a pilot experiment to determine what concentrations of casein and trypsin are best, and what kind of shaking regime is best to ensure the suspension doesnt settle at the bottom.
Control: keep the batch of casein and trypsin the same as you cannot ensure the concentration will be exactly the same in another batch. ...
Apparatus: -
3 test tubes to mix the substrate and enzyme in.
2 syringes (one for casein, one for trypsin)
Test tube rack (to carry out the experiment at room temperature)
105ml of 4% milk Marvel suspension.
Approximate Word count = 3173
Approximate Pages = 12.7
(250 words per page double spaced)
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