Translation Modeling Activity

prusaprinters

Summary continuedThis activity picks up where Transcription leaves off. We will assume that RNA polymerase has already created the mRNA transcript and the mRNA has been processed (addition of the 5’ cap, polyA tail and removal of introns). The mRNA has moved into the cytoplasm of the cell. Translation is ready to occur.Translation is defined as the synthesis of a protein (polypeptide) using information encoded in an mRNA molecule. Three RNA molecules will be used to assemble the protein: Messenger RNA, Transfer RNA and Ribosomal RNA. Messenger RNA (mRNA) has the information for arranging the amino acids in the correct order to make a functional protein. Transfer RNA (tRNA) carries amino acids to the ribosome and Ribosomal RNA (rRNA) makes up the ribosome.The key to deciphering DNA is called a triplet code, in which the sequence of three adjacent DNA nitrogen bases codes for a specific amino acid. Translation of the mRNA occurs in groups of three nitrogenous bases called codons. The order in which the amino acids are put together depends on the sequence of bases in the mRNA. Proteins can consist of as few as 100 or as many as thousands of amino acids.Lesson Plan and ActivityPre-lab Questions1. Where does mRNA come from? What process created it and where in the cell did this occur? 2. How do you know that it is mRNA and not DNA? 3. What process will you be modeling? Where does this process occur? 4. What part of an mRNA nucleotide provides the information to make a protein? 5. Which end of the mRNA strand enters the ribosome first and is attached to the small ribosomal subunit? 6. What are the monomers (building blocks) of proteins?tRNA Charging ProcedureBefore you start translation of the mRNA, you need to load up ALL TWENTY of the tRNAs (green pieces) with the correct amino acids. Take out the tRNA pieces. In the chart below, write the anticodon (the bottom of the tRNA), and then its complementary codon. Look up the codon on the mRNA codon chart and attach the correct amino acid to the tRNA (remember: the amino acid matches with the Codon, not the Anticodon).TranslationTake out one mRNA polynucleotide strand (which can be found in the box labeled mRNAs). Look for the start codon and place it on the ribosome sheet in the proper orientation (AUG on the left). Record the sequence on the line below. mRNA Sequence: Record the number on the back of the mRNA: On the Ribosome The A, P, and E sites are labeled. In the space below, describe what happens at each stage: Initiation: Slide your mRNA into the small ribosomal subunit. Attach the first tRNA-amino acid complex to the mRNA in the P site. Record which tRNA anticodon and accompanying amino acid will attatch first in this P site. ___________________ Elongation: The anticodon of another tRNA base pairs with the mRNA in the A site. Complete this next step using your model. Which tRNA-amino acid complex will attach into the A site at this time? Connect the two amino acids, and detach the first amino acid from the first tRNA, then slide the Ribosome sheet under the mRNA so that the first tRNA is now in the E site and the second tRNA (holding the growing polypeptide chain) is in the P site. Then detach (“Eject”) the first tRNA from the mRNA and remove it from the ribosome. What happens to the tRNA after it leaves at the E site? Continue this process by bringing in the next tRNA to the A site, attaching the amino acid chain to the new amino acid, detaching the chain from the tRNA in the P site and then sliding the ribosome so that the tRNA missing the amino acid is in the E site and ejecting the empty tRNA. Continue until the polypeptide chain is complete. Record your final polypeptide chain’s amino acid sequence:TerminationTermination: 1. Now you have arrived at the codon UGA. What is this codon? In the space below, describe what happens to the ribosome complex, the tRNA molecules and the amino acid chain. 2. Once your (mini-)polypeptide chain is completed, you must disassemble it to create a new one. How does this process mirror what happens in an actual cell? 3. Once you complete your first Polypeptide chain (protein), take out a new mRNA and swap roles with your team members (e.g. if you were in charge of the ribosome, while a partner was in charge of the tRNAs, swap places). Make sure to recharge your tRNA’s with their correct amino acids.Post Lab Questions1. What codon did all mRNA polynucleotide chains start with? What is the significance of this? 2. What ensures that the correct amino acid is brought to the protein in the correct order? 3. What types of bonds hold the anticodon to the codons? What types of bonds hold the amino acids together? How does the strength of these bonds differ? 4. What would happen if the UGU codon was changed to UGC? How would this affect the protein? 5. What would happen if the UGU codon was changed to UGA? How would this affect the protein? 6. What would happen if the UGU codon was changed to UGG? How would this affect the protein? 7. Once the small polypeptide is formed through translation, what happens to it (i.e., what post-translational modifications happen now)? 8. How long did this process of translation take for you and your lab group? Do you think the cell could operate at this rate? Explain. 9. mRNA, tRNA, and ribosomes can be reused over and over. The same protein can be made again if needed, or a new piece of mRNA can be translated. Ribosomes add new amino acids to the polypeptide at a rate of 20 amino acids per second (at 37°C). At this rate, how long would it take to make a small protein such as actin which is 375 amino acids long?Materials NeededThe full classroom set includes ten strands of mRNA, which rotate around to each group of students. Each group receives a set of twenty tRNA, twenty amino acids, one ribosome, one release factor protein, and one "stop" piece. The "stop" piece is representative of the hydrolysis of bonds which occurs as a result of the attachment of a release factor protein when an mRNA strand reaches a stop codon. COSTThe print a full set, which includes the 10 long, yellow mRNA strands and all of the amino acids is $27.61. To find this, I found the cost per part in Simplify 3D, multiplied each part by the number of parts needed for one set, and then added the total cost. In the column all the way to the left I have the part name, next is the number of parts needed for a complete set, and then the cost per part which I found on Simplify 3D. Next to the right is the total mass in grams of the number of parts needed for each piece, which I also found by using simplify 3D. I also found the mass for 6 sets, which is ideal for a classroom. After that, I labeled each piece name a color, so that when two part names had the same color I could add of the costs of both for blue for example. In order to figure out how that translated to each filament color, I had to figure out how many grams were needed for each color. To do this, I used the mass column and added up the mass for each color. Then, I found the  price of each Hatchbox spool and found the price of that. Each Hatchbox is 1 kg, so 1000g. So I multiplied the mass for each color by the price for each spool and then divided that by 1000g to get the price for each color (dimensional analysis). Then, I added up the prices for each of the colors to get the total. The final cost per set is $27.61 and the price for 6 sets is $165.66.