Materials Included In Kit

Safety Precautions

The items used in this activity are considered nonhazardous.

Disposal

All items may be saved and reused for future classes.

Lab Hints

• Enough materials are provided in this kit for 30 students working in pairs or for 15 groups of students. This laboratory activity can reasonably be completed in one 50-minute class period.
• Copies of the Ribosome Master Sheet and Codon Wheel should be made for each student group before class.
• Groups may be given extra mRNA strands to synthesize more polypeptides if time allows.

Teacher Tips

• Consider laminating mRNA strands, tRNA cards and amino acid cards for use in future classes.
• Students should have a basic understanding of monomers and polymers, nucleic acids, transcription and translation before beginning this activity.
• A summary discussion after each step may be worthwhile to be sure of students’ comprehension before moving onto the next step.
• This is an excellent complementary activity to Flinn Scientific’s DNA in Action Kit, Catalog No. FB1223.

Answers to Prelab Questions

1. Define the following terms:

   mRNA—Ribonucleic acid that contains instructions for assembling amino acids into proteins.
   tRNA—Ribonucleic acid that attaches to transfer amino acids and binds to the ribosomes.
   Ribosome—A molecule in the cell on which proteins are assembled.
   Codon—A three-nucleotide sequence on an mRNA molecule representing a specific amino acid.
   Anticodon—tRNA sequences that are complementary to their respective codons on mRNA.
   Translation—Process in which the order of bases in mRNA codes for the order of amino acids in a protein.

2. Which amino acid begins every protein chain? Why?

   Methionine begins every protein chain. The structure of the amino acid methionine (AUG) allows it to go directly to the peptidyl tRNA site to initiate the synthesis of the protein chain.

3. In what part of the cell does translation take place? How does this vary from transcription?

   Translation takes place in the cytoplasm of the cell. Transcription takes place in the nucleus of the cell.

4. Describe, in your own words, the process of protein synthesis.

   Answers will vary. See the student background section and classroom textbook.

Answers to Questions

1. Draw your mRNA strand.
   {11296_Data_Figure_7}
2. Draw the ribosome, methionine/tRNA complex, and the mRNA strand from step 4. Be sure to draw the proper 5′–3′ orientation of the mRNA.
   {11296_Data_Figure_8}
3. Draw the ribosome, mRNA and tRNA/amino acid complexes from step 6.
   {11296_Data_Figure_9}
4. Draw the resulting protein strand from step 12.
   {11296_Data_Figure_10}

Introduction

Piece together the steps of translation by building a polypeptide chain! In the following hands-on activity, students learn the building blocks of a protein and simulate the process of translation by decoding an mRNA molecule.

Concepts

• Protein synthesis
• Transfer RNA
• Codons and anticodons
• Translation
• Amino acids
• Messenger RNA

Background

Every cell in your body contains a copy of your unique DNA. This DNA provides the code for all the proteins in your body. These proteins not only influence how you look on the outside, but also take part in every biological process that occurs within the body. How can one genetic code relay so many different messages?

The code contained in an organism’s genes provides instructions for the synthesis, or building, of a protein. This process from DNA to protein is called gene expression and occurs in two stages—transcription and translation. Transcription is the synthesis of RNA from DNA. This RNA is then processed to result in mRNA (messenger RNA), which carries the genetic message from the DNA to a ribosome. The genetic message is made up of a sequence of codons. Codons contain three bases (A, U, C or G) that code for a specific amino acid. Transcription and RNA processing take place in the nucleus of the cell. Translation occurs in ribosomes outside of the nucleus and is the synthesis of a polypeptide using the information from the mRNA (see Figure 1).

Translation occurs when the message of the mRNA is decoded to form a corresponding protein. Cells use transfer RNA (tRNA) to translate the message from mRNA into a protein. tRNA transfers amino acids (the building blocks of proteins) found in the cytoplasm to a ribosome where translation occurs. One side of the tRNA attaches to an amino acid while the other end contains a specific anti-codon. An anti-codon is a three-nucleotide sequence that is complementary to the codon of the mRNA. In RNA, nucleotide A pairs with U and C pairs with G. For example, the anti-codon for the tRNA molecule carrying the amino acid methionine is UAC, which pairs with the mRNA codon AUG.

Ribosomes coordinate the process of translation, providing an mRNA binding site and three tRNA binding sites. The P-site (peptidyl site) holds the tRNA carrying the growing protein chain. The A-site (aminoacyl site) holds the tRNA carrying the amino acid that will be added next to the chain. The E-site (exit site) is where the tRNA leaves the ribosome.

Protein synthesis always begins with the amino acid methionine. The first AUG codon on the mRNA molecule binds to the P-site on that ribosome. A tRNA with the complementary anti-codon (UAC) brings the amino acid to the site. The tRNA molecule with an anti-codon complementary to the next mRNA codon will bind at the A-site. Once the amino acids are in these sites, a peptide bond forms between the two. Following this, the ribosome moves to the next codon on the mRNA strand, allowing the tRNA molecules in the P-site and A-site to shift. This opens the A-site for the addition of a new amino acid. The bond is then broken between the tRNA in the E-site and its amino acid, allowing the tRNA to leave the ribosome.

The protein chain continues to synthesize in this fashion until the ribosome reaches a stop codon on the mRNA strand. When the stop codon is reached, both the protein chain and the mRNA strand are released and translation is complete.

Experiment Overview

In this activity, models of mRNA, tRNA, amino acids and ribosomes will be used to better understand protein synthesis.

Materials

Prelab Questions

1. Define the following terms:

   mRNA—
   tRNA—
   Ribosome—
   Codon—
   Anti-codon—
   Translation—

2. Which amino acid begins every protein chain? Why?
3. In what part of the cell does translation take place? How does this vary from transcription?

Safety Precautions

The items used in this activity are considered nonhazardous. Follow all classroom guidelines.

Procedure

Part A. Deciphering the Code

1. Obtain a Ribosome Master Sheet, an mRNA strand, a tRNA set (20 cards) and an amino acid set (20 cards).
2. To start, determine the codon that compliments the anticodon on one tRNA card.
   1. Using the codon and the codon wheel, find the associated amino acid. Read the wheel from the inside out. Start from the center of the wheel and locate the first letter of the codon. Move outward, looking for the second and then third letters of the codon. The matching amino acid will be in the outermost shell of the wheel.
   2. Attach the complimentary amino acid to the correct tRNA card using a small piece of tape. Attach near bottom of tRNA card (see Figure 2).
   {11296_Procedure_Figure_2}
3. Repeat step 2 for each of the 20 tRNA cards.

Part B. Translation

1. Locate the methionine (met) amino acid/tRNA complex assembled in step 2. The structure of methionine allows it to go directly to the P-site on the ribosome and start a protein chain. Place the methionine amino acid/tRNA complex on the P-site on the ribosome master sheet. Notice the 3′ to 5′ orientation of the tRNA (see Figure 3).
   {11296_Procedure_Figure_3}
2. Obtain an mRNA strand. Write the base sequence of the mRNA strand in the 5′–3′ direction on the Protein Synthesis Worksheet.
3. Place the mRNA strand on the mRNA site of the ribosome. The mRNA strand should read from 5′ to 3′ from left to right (see Figure 4).
   {11296_Procedure_Figure_4}
4. The first codon of the mRNA strand will be complementary to the anti-codon sequence of the tRNA attached to the methionine amino acid (i.e., the codon AUG is complimentary to the anti-codon UAC). Draw all structures on the Protein Synthesis Worksheet.
5. A codon on the mRNA sequence is now exposed on the A-site on the ribosome. Match up the complementary amino acid/tRNA complex to this mRNA codon.
6. Peptide bond formation now takes place between the methionine amino acid and the amino acid at the A-site. Place a piece of transparent tape from the methionine amino acid to the new amino acid to represent this bond (see Figure 5). Draw all structures on the Protein Synthesis Worksheet.
   {11296_Procedure_Figure_5}
7. Move the ribosome to the right one codon. The amino acid/tRNA complex that was at the A-site will now be at the P-site.
8. Notice that the methionine amino acid/tRNA complex is now at the E-site. This is where the tRNA exits the ribosome. Methionine will remain attached to the second amino acid, forming a polypeptide chain. The tRNA molecule that has broken away from the amino acid enters the cytoplasm in search of another amino acid. Carefully peel off the tape holding the tRNA and amino acid card together.
9. The new mRNA codon at the A-site requires a new amino acid/tRNA complex. Find the complementary amino acid/tRNA complex and place it at the A-site.
10. Place another small piece of transparent tape between the amino acid at the P-site and the new amino acid at the A-site.
11. Move the ribosome to the right once again until the A-site is once again open (see Figure 6).
    {11296_Procedure_Figure_6}
12. Repeat steps 5–11 until the entire mRNA strand has been translated and a complete polypeptide chain has been formed. Draw the final polypeptide sequence on the Protein Synthesis Worksheet and answer the Post-Lab Questions.

Student Worksheet PDF

11296_Student.pdf