Aspartate And Glutamate Lab Report

Eisenthal, R. & M.J. Danson (Eds.) (2002). _Enzyme Assays: A Practical Approach_. United Kingdom: Oxford University Press

We placed the gel into the running chamber, and then we completely covered the gel with TAE. 3 microliters of loading dye was added to each tube; this would help distinguish the enzyme from the gel. As before, we tapped the tube on the table to mix. Then we carefully added each of the four samples into their own wells. A total of 33 microliters of each sample was poured into each well. Afterwards, we attached the positive and negative electrodes to their corresponding terminals on the power supply and gel box. We turned on the power to around 80 volts and waited 45-60 minutes for the loading dye to move down the gel approximately 6-8 cm. Finally, we were able to visualize the DNA in the gel and write down the

There were a couple of problems that our group encountered while conducting this experiment. Since this was our first experiment dealing with enzyme activity, the probability of human error increased. We found it difficult to go through the procedures with undefined roles for our team. The other problem that our team encountered was the getting the cuvettes into the spectrophotometer quickly once the enzyme was introduced to the cuvette.

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

These results shown from this experiment led us to conclude that enzymes work best at certain pH rates. For this particular enzyme, pH 7 worked best. When compared to high levels of pH, the lower levels worked better. The wrong level of pH can denature enzymes; therefore finding the right level is essential. The independent variable was the amount of pH, and the dependent being the rate of oxygen. The results are reliable as they are reinforced by the fact that enzymes typically work best at neutral pH

After the substrate solution was added, five drops of the enzyme were quickly placed in tubes 3, 4 and 5. There were no drops of enzyme added in tubes 1 and 2 and in tube 6 ten drops were added. Once the enzyme solution has been added the tubes were then left to incubate for ten minutes and after five drops of DNSA solution were added to tubes 1 to 6. The tubes were then placed in a hot block at 80-90oC for five minutes. They were then taken out after the five minute period and using a 5 ml pipette, 5 ml of distilled water were added to the 6 tubes and mixed by inversion. Once everything was complete the 6 tubes were then taken to the Milton Roy Company Spectronic 21 and the absorbance of each tube was tested.

Acidosis influences L-glutamate carrier proteins and changes the 3-dimentional structure of the transporter which may finally halt L-glutamate transport (71). Furthermore, acidosis can change the ratio of OH-in to OH- out. This ratio plays an important role in the L-glutamate transporting system on neurons (72) and in turn its disturbances will inhibit L-glutamate (70, 73, 74). Acidosis also stimulates L-glutamate release and this stimulation may participate in the mechanisms by which acidosis inhibits L-glutamate uptake (70, 75). When the stimulation of L-glutamate release and its (re-)uptake inhibition occur simultaneously, it probably leads to an increase in extracellular glutamate which subsequently activates NMDA and AMPA receptors and cause excessive influx of Ca2+ and excitotoxicity (69, 76) Consequently, these NMDA receptors trigger more changes in pHi which seems to be toward acidification. Such a shift results from increases in intracellular sodium and extracellular potassium which is mediated by NMDA receptor activation and subsequently stimulates glycolysis in neurons (77). The mentioned pHi acidification is accompanied by the increased production of lactate whose pathological concentrations may play a role in NMDA receptor-induced neuronal injury or death and neurodegeneration (77-80).

According to the Tufts University Health and Nutrition letter the FDA supports the safety of the toxicology of Aspartame. It has been stated that "The FDA recently rejected two citizen petitions calling for an aspartame

The second neurotransmitter family includes amino acids, compounds that contain both an amino group (NH2) and a carboxylic acid group (COOH) and which are also the building blocks of peptides and proteins. The amino acids known to serve as neurotransmitters are glycine, glutamic and aspartic acids, all present in all proteins, and gamma-amino butyric acid (GABA), produced only in brain neurons. Glutamic acid and GABA are the most abundant neurotransmitters within the central nervous system, particularly in the cerebral cortex; glutamic acid tends to be excitatory and GABA inhibitory. Aspartic acid and glycine subserve these functions in the spinal cord (Cooper, Bloom, and Roth 1996).

The biggest problem my group experienced was that as we moved into our second round of trails, we noticed consistently higher reaction times for the same concentration of enzyme. There are two possible explanations for this. The first is that we continued to use the same container of hydrogen peroxide every trail. The peroxide may have become diluted, reducing the rate at which the reaction could happen. This became apparent when we got to the lowest concentration trails. We were reasonably sure that these would both take a long time, so we did both trails simultaneously in two different containers: one old, one new. The trail conducted in the container of fresh peroxide finished in just over six minutes, however the trail conducted in the used

The purpose of this experiment was to record catalase enzyme activity with different temperatures and substrate concentrations. It was hypothesized that, until all active sites were bound, as the substrate concentration increased, the reaction rate would increase. The first experiment consisted of five different substrate concentrations, 0.8%, 0.4%, 0.2%, 0.1%, and 0% H2O2. The second experiment was completed using 0.8% substrate concentration and four different temperatures of enzymes ranging from cold to boiled. It was hypothesized that as the temperature increased, the reaction rate would increase. This would occur until the enzyme was denatured. The results from the two experiments show that the more substrate concentration,

Neurotransmitters are endogenous chemical compounds that transmit neural signals from one neuron to another. Many neurotransmitters are amino acids, such as glutamate, glycine and GABA, or biogenic amines, such as dopamine and serotonin, or even peptides and proteins, such as somatostatin and substance P {Snyder 1979}. Binding of neurotransmitters may either inhibit or excite the postsynaptic neurons. Among the numerous neurotransmitters, glutamate is the major excitatory amino acid neurotransmitter in mammalian neural systems {Cotman 1986}. The first genetically encoded neurotransmitter for glutamate was reported in 2005 {Okumoto, 2005 #292}. Upon binding to glutamate, the indicator converts the

In this lab or experiment, the aim was to determine the following factors of enzymes: (1) the effects of enzymes concentration the catalytic rate or the rate of the reaction, (2) the effects of pH on a particular enzyme, an enzyme known and referred throughout this experiment as ALP (alkaline phosphate enzyme) and lastly (3) the effects of various temperatures on the reaction or catalytic rate. Throughout the experiment 8 separate cuvettes and tubes are mixed with various solutions (labeled as tables 1,3 & 4 in the apparatus/materials sections of the lab) and tested for the effects of the factors mentioned above (concentration, pH and temperature). The tubes labeled 1-4 are tested for pH with pH paper and by spectrophotometer, cuvettes 1a-4a was tested for concentration and cuvettes labeled 1b-4b was tested for temperature in four different atmospheric conditions (4ºC, 23ºC, 32ºC and 60ºC) to see how the enzyme solution was affected by the various conditions. After carrying out the procedures the results showed that the experiment followed the theory for the most part, which is that all the factors work best at its optimum level. So, the optimum pH that the enzymes reacted at was a pH of 7 (neutral), the optimum temperature that the reactions occurs with the enzymes is a temperature of 4ºC or

In the brain there is an excitory and inhibitory input of neurotransmitters. GABA is the main inhibitory input of neurotransmitters and glutamate is the main excitory input of neurotransmitters. Glutamate transmits chemical signals from neuron to neuron. It is critical for learning and remembering. [1] Glutamate, as it is lethal in high concentrations to nerve cells, is stored in neurons as glutamine in order to keep the neurons healthy and to prevent the stimulation of seizures. [2] Glutamate is synthesised one of two ways, it can either be converted from glutamine to glutamate with the assistance of the enzyme glutaminase, or by transaminating 2-oxoglutarate. [3]

The independent variable in this investigation is pH. Each individual enzyme has it’s own pH characteristic. This is because the hydrogen and ionic bonds between –NH2 and –COOH groups of the polypeptides that make up the enzyme, fix the exact arrangement of the active site of an enzyme. It is crucial to be aware of how even small changes in the