Tyrosinase Lab Report

Enzymes are biological catalysts, which means it decreases activation energy in reactions. The lower activation energy in a reaction, the faster the reaction rate. Many enzymes alter their shape when they bind to the activation site. This is called induced fit, meaning for the enzyme to work to its full potential it has to change shape to binding substrate. The location of enzyme’s activation site is on the surface of the enzyme, where the binding of substrates take place. Enzyme activity can be influenced by a variety of environmental factors. If the concentration of enzyme is low, and there is a great deal of substrate, then increasing enzyme concentration results in more molecules available to convert substrates to products. Thus, increasing enzyme concentration can increase reaction rate. If substrate concentrations are low, and many of the existing enzymes are idle because of a lack of substrate, then adding enzyme will have no effect on reaction rate. Enzyme concentration affects the enzyme activity, because the more enzyme concentration the faster the reaction rate, until it hits it’s limiting factor. When substrate concentration is increased, it also increases rate of reaction. Temperature plays an important

However, temperature difference can affect the experiment. To prevent this, I will conduct the experiment at the same time of day if more than one day is needed. This is because it is generally colder in the mornings and evening than the afternoon.

[An active site can be altered by a non-competitive enzyme which encircles the enzyme and alters the shape of the active site which could be very dangerous.]

Background and Introduction: Enzymes are proteins that process substrates, which is the chemical molecule that enzymes work on to make products. Enzyme purpose is to increase the rate of activity and speed up chemical reaction in a form of biological catalysts. The enzymes specialize in lowering the activation energy to start the process. Enzymes are very specific in their process, each substrate is designed to fit with a specific substrate and the enzyme and substrate link at the active site. The binding of a substrate to the active site of an enzyme is a very specific interaction. Active sites are clefts or grooves on the surface of an enzyme, usually composed of amino acids from different parts of the polypeptide chain that are brought together in the tertiary structure of the folded protein. Substrates initially bind to the active site by noncovalent interactions, including hydrogen bonds, ionic bonds, and hydrophobic interactions. Once a substrate is bound to the active site of an enzyme, multiple mechanisms can accelerate its conversion to the product of the reaction. But sometimes, these enzymes fail or succeed to increase the rate of action because of various factors that limit the action. These factors can be known as temperature, acidity levels (pH), enzyme and/or substrate concentration, etc. In this experiment, it will be tested how much of an effect

J. Moldovan & B. Nilson, (2010), Lab 4 – Enzyme Kinetics, UBCO BIOL/BIOC 393, UBC Vista accessed Monday, November 8th, 2010.

Enzymes have things called ‘Active Sites’ this the spot where substrates bind and chemical reaction takes place. Each enzyme has a different shape, this allows the enzyme to receive only one type of

From the stock substrate solution of 2.5 mM, each group serially diluted at least one different substrate concentration for a total of four different substrate concentrations to be investigated: 1.25 mM, 1.0 mM, 0.75 mM, 0.25 mM. The enzyme concentration was kept constant at 2.0 mM while experimenting on the affect of varying enzyme concentration on the rate and product formation of ONP. Enough 2.0 mM enzyme solution was prepared in the previous part of the project to supply this assay. Using similar procedure to collect absorbance data as the first part, 0.5 mL of 2.0 mM enzyme concentration was placed into the cuvette and used to calibrate the spectrometer at 420 nm. Data was then started, with the immediate addition of 0.5 mL of varying substrate concentrations. Each varying substrate concentration was split between the team and run for a total of 10 minutes, with the exception of the 1.25 mM run. Upon completion, data from each varying substrate concentration was copied to a single Excel sheet and used to produce an absorbance vs. time graph, product formation vs. time graph, Michaelis Menten plot, and Lineweaver-Birk plot. This analysis was used to calculate the V0,Vmax, and Km for β-Galactosidase

Enzymes are proteins that catalyze (speed up) biological reactions in an organism by lowering the activation energy of a reaction. They do this by either straining the bonds in a molecule so that is easier to break up or by placing separate molecules/elements close to each other so that bonds are formed. Enzyme activity is influenced by an array of different factors such as enzyme concentration, substrate concentration, temperature, pH and inhibitor concentration. All of these affect the rate of reactions of enzymes and some such as temperature, inhibitors and pH can under circumstances cause enzymes to become permanently affected. Catalase is an enzyme found in almost all organisms on earth exposed to Oxygen.

However, the rate of reaction only increases for a certain period of time until there is lesser substrate molecules than the enzyme molecules. The increase of enzyme concentration does not have effect if there are lesser substrate molecules than enzyme molecules initially.

This is because the more neutral the pH the faster enzymes can catalyse reactions. If there is an excess of H⁺ or OH⁻ ions then the shape of the protein alters, thus slowing down catalysis. The graph shows this as pH7 is neutral and has the highest reaction rates, while pH3 and pH11, acidic and basic, respectively, both have fairly low reaction rates. pH5 and pH9 are both in the middle because they are slightly acidic, so they have a lower reaction rate than pH7, but a higher reaction rate than pH3 and pH11. Describe the relationship between substrate concentration and the initial reaction rate of an enzyme-catalyzed reaction.

Organisms cannot depend solely on spontaneous reactions for the production of materials because they occur slowly and are not responsive to the organism's needs (Martineau, Dean, et al, Laboratory Manual, 43). In order to speed up the reaction process, cells use enzymes as biological catalysts. Enzymes are able to speed up the reaction through lowering activation energy. Additionally, enzymes facilitate reactions without being consumed (manual,43). Each enzyme acts on a specific molecule or set of molecules referred to as the enzyme's substrate and the results of this reaction are called products (manual 43). As a result, enzymes promote a reaction so that substrates are converted into products on a faster pace (manual 43). Most enzymes are proteins whose structure is determined by its sequence of its amino acids. Enzymes are designed to function the best under physiological conditions of PH and temperature. Any change of these variables that change the conformation of the enzyme will destroy or enhance enzyme activity(manual, 43).

An enzyme is a catalyst. Catalysts are known for speeding up the rate of reactions by lowering the activation energy of the biochemical reaction. (Reece et al., 2011)

reaction mechanism. Therefore, if an enzyme has a high KM value, then it has a relatively low affinity for its substrate since it requires a higher substrate concentration to reach its vmax. In contrast, a low KM value corresponds to a high substrate affinity. The vmax value corresponds to the rate of reaction at which the enzyme is completely saturated with substrate. Therefore, this vmax value reflects how fast an enzyme can catalyze a given reaction.

When unlocking a door, only a certain shape and size of a key will fit into the lock. This is similar with and enzyme and a substrate. Only a specific shape and size substrate will fit into the enzyme’s active site (figure 1B). The induced fit model explains how an enzyme’s active site many change after the enzyme and the substrate are combined (figure

The purpose of this lab report is to investigate the effect of substrate concentration on enzyme activity as tested with the enzyme catalase and the substrate hydrogen peroxide at several concentrations to produce oxygen. It was assumed that an increase in hydrogen peroxide concentration would decrease the amount of time the paper circle with the enzyme catalase present on it, sowing an increase in enzyme activity. Therefore it can be hypothesised that there would be an effect on catalase activity from the increase in hydrogen peroxide concentration measured in time for the paper circle to ride to the top of the solution.