Ashiana, Taylor and Raffi

April 17th 2019

           The Effect that Temperature has on Glucose Concentration

Introduction: The purpose of this experiment is to better comprehend the effect of temperature on enzyme reaction rate. It is also to come to the understanding that lactose is a disaccharide made up of one glucose and one galactose. When the temperature of the substrate is increased, the lactase drops will act as a catalyst at a faster rate and there will be more glucose measured by the test strips. This is because when the temperature of substrate is increased, the reaction rate speeds up. Heat is a form of energy, when the energy is increased the particles move faster increasing the rate in which they bump into each other. Hypothesis: If the temperature of the substrate goes up, then their will be a higher glucose concentration.

Materials:

• 1 2% cream box (we will provide)
• Lactase drops
• 6 Glucose test strips
• 6 Test tubes
• 1 Test tube Rack
• 1 Kettle
• 250mL graduated cylinder
• Water
• Ice
• 250 mL Beaker
• Thermometer

Procedure:

1. Measure and pour 15mL creme in the 6 test tubes on the test tube rack.
2. Fill up the beaker with water and 15 ice cubes. When the water reaches below 0, measure 15mL into the graduated cylinder and pour it into the 1st test tube on the rack that holds the creme. The mixture should reach 5 degrees celsius right before the glucose strip is inserted. 
3. Mix in 5 lactase drops.
4. Wait 15 minutes, check the that the mixture has reached the desired temperature, insert glucose test strips.
5. Wait until the beaker with the water has warmed up to around 5 degrees celsius. Measure 15mL into the graduated cylinder, pour it into the 2nd test tube on the rack that holds the creme. The mixture should reach 10 degrees celsius right before the glucose strip is inserted.
6. Repeat steps 3-4.
7. Wait until the beaker with the water has warmed up to around 8 degrees celsius. Measure 15mL into the graduated cylinder, pour it into the 3rd test tube on the rack that holds the creme. The mixture should reach 15 degrees celsius right before the glucose strip is inserted.
8. Repeat steps 3-4.
9. Empty the beaker and dry it out.
10. Pour room temperature tap water into the beaker. Measure 15mL into the graduated cylinder and pour it into the 4th test tube on the rack that holds the creme. The mixture should reach 20 degrees celsius right before the glucose strip is inserted.
11. Repeat steps 3-4.
12. Empty the beaker and dry it out.
13. Boil water in the kettle, pour it into the beaker and let it cool down to around 40 degrees. Measure 15mL into the graduated cylinder and pour it into the last test tube on the rack that holds the creme. The mixture should reach 30 degrees celsius right before the glucose strip is inserted.
14. Repeat steps 3-4.
15. Because the water in the beaker has been sitting, it should have cooled to 35 degrees. Measure 15mL into the graduated cylinder and pour it into the 5th test tube on the rack that holds the creme. The mixture should reach 25 degrees celsius right before the glucose strip is inserted.

Note that sometimes trial and error is needed to receive the desired variables.

The independent variable in this experiment was the temperature that the creme was. The dependant variable was the concentration of glucose that was resulted from the temperature. The controls in this experiment were the amount of substrate (creme) used, the concentration of the enzyme (5 drops of the lactase drops) the type of creme used and the amount of water per test tube.

Results:

Data Analysis: It is interesting to compare the lowest temperature and the highest temperature because it followed the hypothesis and clearly differed in the amount of glucose formed. The reaction happened a lot more when the creme was 30 degrees in comparison to when it was 5 degrees. However, the middle temperatures from 10-25 all had the same amount of glucose concentration, which still follow the hypothesis because they were in between the high and low. Except for when it was 15 degrees, the glucose concentration was unexplainably a lot higher.  If you look at the photo you can see how the 20 and 25 degree corresponding glucose test strips have darker edges around their light brown squares in comparison to the 10 degree which also follows the hypothesis. This overall demonstrates an increase in glucose concentration with the addition of heat.

Conclusion: In conclusion, overall the hypothesis was confirmed as their was generally an increase in glucose concentration with the addition of heat. This confirms that with the increase in temperature their is an increase in energy in the substrate that allows the enzymes and substrate to bump into each other more often, therefore increasing the rate of the reaction. Because their was more glucose when the temperature of the substrate was 30 degrees in comparison to when the temperature was 5 degrees, one can infer that lactose decomposes into glucose and galactose more efficiently with the addition of heat. When the substrate was 15 degrees there was clearly a mistake made as the hypothesis was refuted only at that specific temperature and therefore the experiment should be done again to further confirm the hypothesis.

Errors/Improvements: It was very difficult to maintain a specific temperature after adding lactase drops and waiting 15 minutes to put the glucose test strips in. The temperature continuously fluctuated and trial and error was done in adding different temperatures of water in after the 15mL of water initially added. This definitely had an effect on the results. When the substrate was 15 degrees, because it was the closest to room temperature with the addition of creme, there was no need for the addition of water. Therefore, the proportion of substrate to enzyme was more even it is possible that it increased the reaction rate. Because when the substrate was 20 degrees it required the input of warm water to keep up the temperature, there was more substrate in comparison to the enzyme so enzyme concentration was less. In order to improve this experiment there will need to be refinements made in the experiment. It will be more beneficial to have full knowledge on how fast the temperature will fluctuate so that it can be timed out that it will reach the exact temperature wanted in the 15 minutes that the enzyme needs to mix with the substrate, prior to adding the glucose test strips. The procedure will differ for every environment that the procedure is executed in, in order to maintain temperatures.

The fact that the amounts of substrate varied in each test tube is a huge error as it will effect the enzyme concentration which was supposed to be a control group. However, this was required in the experiment to maintain the temperatures desired.

RNA Transcription Model

1. How is mRNA different then DNA

mRNA differs from DNA in the sense that it uses the ribose sugar in lieu of deoxyribose. It is shorter then DNA which allows it to exit from the nuclear pore – something that DNA is not capable of. It has one strand instead of two, making it a single backbone. It does not have an double helix shape. mRNA substitutes the pyrimidine uracil for the pyrimidine thymine, meaning that all the Adenines on DNA are paired with Uracil on RNA.

1. Describe the process of transcription

This whole process is essential because DNA carries the information for protein synthesis but is too large and vital to the nucleus to exit. Protein synthesis must occur in the cytoplasm of the cell where the ribosomes, or protein factories are located. mRNA is derived form the process of transcription in order to be able to exit the nuclear pores with DNA’s information and produce codons to be read by the ribosomes later in translation for the construction of proteins.

1. Unwinding and unzipping of DNA

DNA helix shape unwinds. The unzipping of DNA only happens at the desired location of the gene for specific information. Exposing instructions to build the gene that is running low only. The whole 85 million long nucleotide strand does not unzip. The sense strand is the strand that RNA has it’s information transcribed from. The sense strand is the strand that carries the information for protein synthesis. The nonsense strand is useless in this process  as it  would spit out non readable info if transcribed, it bonds as a compliment to the sense strand on DNA. 

2. Complimentary based pairing with DNA

Hydrogen bonds are formed between the complimentary based pairs. RNA does not posses the pyrimidine thymine and instead the pyrimidine uracil is bonded to adenine. Guanine and Cytosine remain pairs. The  RNA strand (in red) is base pairing with the nitrogen bases on the sense strand (in blue). This is essential so that the information is copied precisely. 

3.The RNA polymerase Enzyme, represented by a fuzzy peach in this picture facilitates the bonding between the nearby nucleotides. There is a sugar & phosphate covalent bond. 

4. Separation from DNA

RNA is separated from DNA, and DNA re-zips and returns to it’s original double helix shape.

Its’ the messenger RNA (mRNA) that is now developed by the process of replication of the information the gene on DNA possessed. It is checked and refined to ensure that the introns (unnecessary sections) are removed as well as to ensure that it is capped at the ends. It then can exit through the nuclear pore.

1. How did today’s activity do a good job of modelling the process of RNA transcription? In what ways was our model inaccurate.

Accuracies:

• We modelled only one strand for RNA and two strands in a helix shape for DNA.
• We used a red strand to distinguish ribose sugar and blue for deoxyribose sugar.
• We used a brown bead for Uracil the pyrimidine that binds with Adenine in RNA and a Blue beed for Thymine the pyrimidine that binds with Adenine in DNA.
• We modelled RNA polymerase using a fuzzy peach showing how it bonds the nucleotides in a row.

Inaccuracies:

• We demonstrated the transcription on our entire model as it was only 18 nucleotides long however generally the RNA strand should open up, as we demonstrated in the unwinding and unzipping process and then the RNA strand should enter like so:

• It’s inaccurate how the sense strand and the RNA strand were connected with no sign of the complimentary strand as the above photo shows that the RNA strand should enter inside the double bond of DNA and even thought it has unzipped the RNA strand should not completely replaced the complimentary strand, as the model showed  in step 2:

• We did not demonstrate the process of the mRNA being refined before modelling translation down below.

Protein Synthesis Model

1. Describe the process of translation: initiation, elongation and termination.

Prior to Translation, the process of transcription (as shown above) must occur in order for mRNA to be derived from the DNA sense strand. Here is a short summary:

1. Initiation

The mRNA released from the nuclear pores and connected by transcription bonds with a small ribosome subunit in the cytoplasm. The small and large ribosome subunits then bond together. Ribosome starts to read the mRNA that has bonded to it and waits for the start codon AUG. mRNA start codon is AUG, codons produced by mRNA are complimented by anticodons produced by tRNA. There are 64 3 letter codons and from these 64: 20 amino acids can be formed. There are two binding sites in the ribosome, one is the P site and one is the A site.  This means that a ribosome can contain two tRNA’s at a time. The initial tRNA binds to the P site, and it contains one amino acid  Methionine the amino acid formed by the start codon AUG.

2. Elongation

Another tRNA binds in the A site:

The amino acid from the initial tRNA moves to bond with the amino acid in the new tRNA in the A site. The initial tRNA is released.

This opens up a space and the remaining tRNA is moved to the P site. A new tRNA binds in the A site.

The process continues as the amino acids from the tRNA In the P site are moved to the A site so that the tRNA in the P site can be released. Freeing up a space so that the tRNA in the A site can move over to the P site, making room for the a new tRNA.

3. Termination

A stop codon is read on the mRNA  – UAG, UGA or UAA. There is no amino acid that is produced from any of these stop codons which means that the process is terminated. A release factor binds in the A site, in lieu of a new tRNA, causing the breakage of the bond between the tRNA and the P site (the final tRNA). The subunits of the ribosome disconnect and the polypeptide chain of amino acids – the protein, is released.

1. How did today’s activity do a good job of modelling the process of translation? In what ways was our model inaccurate.

Accuracies: 

•  The start codon was modelled, as well as the anticodon that complimented it, as shown in the initiation process above. Methionine was accurately the first protein.
• The two sites (A and P) were shown as well as the process of the new tRNA bind in the A site and the proteins transfer from the older tRNA in the P site with the newest protein in the A site.
• The process of elongation was modelled nicely.

Inaccuracies:

• We did not show the initial smaller ribosome subunit bonding with only the initial tRNA. We showed the initial tRNA bonding with the entire ribosomal unit when in the textbook it explains that the initial tRNA bonds with  only the smaller ribosomal unit and then bonding afterwards with the larger ribosomal subunit for completion of the ribosome.

• We did not model the stop codon UAG, UGA or UAA.
• We did not model a release factor bonding in the A site in order for the polypeptide to dissociate and get released.
• We did not model the separation of the ribosomal subunits just as we did not model when they bind together. The separation occurs with the release of the polypeptide.
• Finally, we could have modelled it better, by showing the release of the tRNA each time a little more clearer then simply having it disappear. We could have shown it outside the ribosomal unit.