Mitochondrial Enzyme Lab Report

The purpose of this lab experiment was to determine the relationship between temperature and the rate of enzymatic activity in yeast cells. In the lab, the temperature was the independent variable. The temperatures consisted of 6°C, 24°C, 34°C, 46°C. The dependent variable in the experiment was the rate of enzymatic activity in yeast cells. The temperatures were tested by using a LabQuest and pressure probe that tested pressure inside the plastic test tube. LabQuest graphed the data and created a line of best fit that was used to determine the slope of the graph. The slope of the graph represents the rate of enzymatic activity. The slope was found for each temperature in 2 different trials. Then, the rate of enzyme activity (kPa/sec) for each temperature in the 2 trials were averaged. These averages were used to develop a graph that shows the relationship between temperature and the rate of enzyme activity. According to the results of the experiment, as the temperature increases, the rate of enzymatic activity decreases. Each enzyme has its own optimal temperature in which it can function efficiently.

To test the effect of temperature, place 4, clean test tubes in a test tube rack and label them “T 0-5,” “T 20-25,” “T 30-35,” and, “T 50-55.” Add 3 mL of 3% H2O2 and 3 mL of water to each test tube. To measure enzyme activity at 0-5°C, prepare a water bath with a temperature range of 0-5°C in a 600 mL beaker. Place Test Tube T 0-5 in the water bath and record the temperature in Table 4. Add 2 drops of the enzyme solution to the test tube, and connect the free end of the plastic tubing to the connector in the stopper. Click the “Collect” button to collect data, and

At the end of three minutes the absorbance level was only 0.234. The hot temperature seems to have slowed the enzyme’s ability. The other group’s ice bath reading started at a low 1.326 it then grew rapidly over thirty seconds to the 2.4 range that the room temperature was also at. The cold temperature seemed to slow the enzymatic activity. The difference between the cold and room temperatures can be observed in graph 1

The first hypothesis was unsupported as Tube 4 (positive control) had the most succinate (.2mL) but ended with the lowest % transmittance out of the three sample we were examining (Tubes 2-4). We believe this discrepancy to be due to the variability of the mitochondrial solution. If the Tube 4 mitochondrial suspension had less concentrated enzyme than the solution for Tube 3 and Tube 2 then Tube 4 would be biologically incapable of matching their rate of reduction. It's worth mentioning that Tube 2 and 3 had a %T change of 23.1% and 23.2% respectively while Tube 4 exhibited a change of only 20.4%; further supporting the idea that Tube 4, despite its increase in succinate concentration, wa lacking in other enzymes/proteins necessary for the

It achieves this my measuring the mitochondrial enzyme activity and as such is used to determine the cytotoxicity of potential medical drugs. It measures the reduction of yellow 3-(4,5-dimethythiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) by mitochondrial succinate dehydrogenase. The MTT starts as yellow but as it enters the cells and passes into the mitochondria where it is reduced and changes in colour to a dark purple formazan product. The cells are then solubilised with an organic solvent and released, solubilised formazan reagent is measured using a spectrometer (Cree, 2011). Some errors that may occur while using this assay are that uneven evaporation of culture fluid in the wells may cause erroneous results. The MTT solution is stable for 6 months when stored at 0°C but at 2-8°C for more than 2 weeks may cause decomposition and erroneous results may occur. Microbial contamination will contribute to cleavage of MTT and the formation of MTT formazan, causing erroneous results (Sigma Aldrich,

Enzymes only bind to specific substrates. Even a small change in the structure of a molecule can make an enzyme inefficient. Test tube 1 was the positive control of the experiment. Whereas, test tube 2 and 3 were the negative controls of the experiment. Results for the effect of temperature on enzyme showed that the test tube which was heated ate 80°C showed no color change, this means that if an enzyme is heated above its optimum temperature enzyme activity decreases, and the enzyme is denatured.

In Experiment One, the data signifies that the velocity of the enzyme increased as the concentration of the substrate increased. However, the data from the second experiment draws inconclusive. The qualitative observations of the color change were a clear sign to start the experiment over again due to possible contamination, and that was not done. This could possibly be a reason why the data shown in the effects of temperature is so skewed. While temperature should rapidly increase the rate of enzyme activity until a certain point at which the enzyme denatures, the data shows denaturation of all enzymes besides Tubes 3 and 4, the color changing solutions in both parts of Experiment Two.

Further research alludes to the fact that this may be the first study which demonstrates increased mitochondrial function as caused by the bacterium’s polysaccharide outer capsule. Researchers are in the process of synthesizing drugs that could potentially protect the mitochondria and greatly diminish cell death, as well as bacterial replication by blocking the effects of the outer capsule. This particular group intends on advancing their study of drugs to mice and to observe its

The hub of energy metabolism, the mitochondrion, is found in virtually all eukaryotic cells, with the exception being erythrocytes. The mitochondrion generates cellular energy in the form of adenosine triphosphate (ATP), mostly by means of the oxidative phosphorylation (OXPHOS) system that is located in the inner mitochondrial membrane. The respiratory chain (CI-CIV) and ATP synthase (CV) is collectively known as the OXPHOS system, encoded by both nuclear DNA (nDNA) and mitochondrial DNA (mtDNA). The number of mitochondria per cell, ranging from hundreds to thousands, is controlled by the energy requirements of specific tissues with the greatest abundance of mitochondria found in metabolic active tissue (Pieczenik and Neustadt, 2007). Mitochondrial disease is caused when there is a defect in any of the numerous mitochondrial pathways, due to spontaneous or inherited mutations. Respiratory chain deficiencies (RCDs) are the largest subgroup of mitochondrial disease and occur when one of the four respiratory chain complexes become impaired. RCDs are considered to be one of the most common

The biochemical process at work is the cause of mitochondrion, a membrane-bound organelle found in the cytoplasm of

The purpose of this experiement is to determine the effect of temperature on the rate of enzyme

Conversely, in sample tube 4, DPIP and succinate were present, however there was no sign of mitochondria. Thus the percent transmittance was extremely constant and essentially lessened for sample 4. According to the initial reaction rates table, samples presenting a

Temperature has a negative and positive effect on enzymes. As the temperature increases from 0 to 40 degrees (See Fig 2) the movement of the enzyme and substrate quicken and will bind more often.But, as the temperature increase from 40 degrees the enzyme and substrate slow and cannot bind as quick and therefore at 63 degrees production stops.

To determine how different temperatures and how they can directly affect the outcome of enzyme activity, we will need to look at the actual temperatures used in the study as well as the specific type of enzyme used. In remembering that enzymes; Proteins speed up the rates of chemical reactions in the types of living organisms used in the lab experiment this report is based upon. With our groups’ hypotheses, including prediction: The determination factor being can a higher temperature decrease the enzyme activity? In this study we as a lab group needed to be able to form conclusions with knowing at what temperatures did we see the results?

A team of researchers from Boston University, supervised by Thomas Tien, studied the mitochondrial function