Materials Included In Kit

Additional Materials Required

Prelab Preparation

Cut one 10-meter piece of string for each student.

Safety Precautions

Remind students to wash their hands thoroughly with soap and water before leaving the laboratory.

Lab Hints

• Enough materials are provided in this kit for 30 students to make each virus model. Both parts of this activity can reasonably be completed in one 50-minute class period. The pre-laboratory assignment may be completed before coming to lab.
• Students may have difficulty with folding the icosahedral shape on the lines. Using the edge of a ruler to burnish the creases makes this task easier and produces more accurate folds.

Further Extensions

• Extend the activity by having students create a bacteriophage using the icosahedron and wire.
• Assign other viruses to the students and allow them to research and create their own models of the assigned virus.

Answers to Prelab Questions

1. One nucleotide of RNA or DNA is approximately 0.34 nanometers long. One millimeter equals 1,000,000 nm. If the genetic sequence for HIV is 9749 nucleotides, how long is the genome for HIV in nanometers? In millimeters?

   9749 nucleotides x 0.34 nm/nucleotide = 3400 nm
   3400 nm x 1 mm/1,000,000 nm = 0.0034 mm

2. A single HIV capsid is 120 nanometers (nm) in diameter. In order to make a model of HIV the capsid is scaled up to 120 mm. How long would a scaled model of the genome be? Hint: Refer to the Background section.
   {11107_PreLabAnswers_Equation_1}

   0.00012 mmx = 0.36
   X = 3400 mm (or 3.4 meters)

Answers to Questions

Part A. Polio Virus

1. Diameter of model polio capsid in mm.

   60 mm

2. Length of live polio capsid in nm.
   {11107_Answers_Equation_2}
3. Length of model polio genome in mm.

   Live polio capsid = 30 nm in diameter
   Model polio capsid = 60 mm in diameter

   {11107_Answers_Equation_3}

   X = 5000 mm

Part B. Tobacco Mosaic Virus

4. Length of live TMV genome in nm.
   {11107_Answers_Equation_4}
5. Length of model TMV genome in mm.
   {11107_Answers_Equation_5}

   X = 440 mm

6. Length of full-size TMV model genome in mm.
   {11107_Answers_Equation_6}

   X = 45,000 mm (or 4.5 meters)

Post-Lab Analysis

7. Why is an enveloped virus more difficult to treat or vaccinate against than a virus without a lipid layer?

   The immune system of the host is delayed in its response because of the camouflauge provided by the envelope.

References

Protein Data Bank, Tobacco Mosaic Virus. http://www.pdb.org/pdb/101/motm.do?momID=109 (accessed July 2011).

Riedel, S. Edward Jenner and the history of smallpox and vaccination. Baylor University Medical Center [Online] 2005, http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1200696/ (accessed July 2011).

Introduction

Everyone has been infected by a virus at some point in his or her life. Explore the biological niche of viruses by studying their shape and composition.

Concepts

• Viruses
• RNA and DNA
• Capsid shape
• Envelope

Background

Viruses seem very simple. They are composed of a chain of genetic material and one or more proteins. The proteins form a shell around the genetic material. This protein shell is called the capsid. The genetic material can be either RNA or DNA depending on the type of virus. The entire virus—the capsid plus the nucleic acid strand—is called a viron. Some viruses have a third feature, a lipid layer that it takes from the host cell as it exits the cell. This layer is called the envelope. Not all viruses have an envelope.

Viruses can be thought of as parasites in that they must use the cell processes of a host to replicate. Since viruses infect cells they must be much smaller than the cell itself. Some viruses are actually larger than the smallest bacteria but they are always smaller than the host cells they infect. Different types of viruses infect specific hosts. Some viruses only infect elm trees, others only infect specific bacteria, and a few are known to only infect humans.

Some viruses contain DNA while others contain only RNA. This is one of the major differences between types of viruses. The simplest RNA virus contains a single strand of RNA that is only 1000 bases long. This extremely short genetic sequence codes for two proteins—that’s it. The largest virus genome contains 1.2 million DNA bases that code for more than 1000 proteins. If the virus contains RNA, the genetic strand is typically decoded within the cytoplasm of the host cell. If the virus contains DNA, the genetic strand is typically decoded within the nucleus of the cell. There are exceptions though. Smallpox is one of a small number of double-stranded DNA viruses that replicate within the cytoplasm.

In general, the larger the genetic strand in a virus the greater the number of proteins the strand encodes and the larger the physical size of the virus. Size varies greatly but there are only three different shape categories. The most common shape of viruses is the icosahedron, a shape that has a soccer ball–like symmetry made of at least 20 faces that are identical isosceles triangles (see Figure 1).

The second most common shape is the helix, a spiral shape (see Figure 2). The third shape is called complex. This is a catch-all category that includes the spaceship-like bacteriophage (see Figure 3a) and the brick-shaped smallpox virus (see Figure 3b). There is no correlation between shape and type of genetic material. DNA viruses can be any of the three shape categories as can RNA viruses. The first virus discovered was the helix-shaped Tobacco Mosaic Virus (TMV). Although first isolated from tobacco leaves this virus remains important as it also infects tomatoes, peppers, and other plants. TMV was first isolated in 1892 and first visualized in 1939 using an electron microscope. The helical capsid is composed of one protein repeated 2130 times so that it forms a hollow tube in the middle. TMV is an RNA-virus containing a single strand of RNA that is 6400 bases long and fits into the inside of the tube, where it is bonded to the helix of proteins. The RNA codes for four proteins: one is the capsid protein, two assist in replicating the RNA strand, and the fourth protein aids in transporting the virus to the next host cell. Genomic replication and protein synthesis take place in the cytoplasm of a host plant cell. Thousands of new viruses are produced within one host cell. Two layers of the capsid form before a completed RNA strand binds to the inside of the capsid. Once the building process begins it proceeds quickly until the viron is complete. TMV will actually self-assemble if capsid proteins and complete RNA strands are mixed together in a lab. Once the structure is completely formed, the TMV virus is extremely stable. Heat and dehydration do not destroy its virulence. That’s amazing considering its small size and simplicity. TMV particles are 300 nm long and 18 nm in diameter. The size of viruses is usually expressed in nanometers (nm), where one millimeter is equivalent to one million nanometers!

Many viruses that infect people have an icosahedral shape. Just as with the helical viruses, icosahedral viruses may contain either RNA or DNA depending on the specific virus. Also, like the helix capsid, the icosahedron capsid is composed of repeating proteins. Whereas the TMV helix is composed of one protein, the simplest icosahedron virus is composed of four proteins. An example of a simple four-protein, icosahedral virus relevant to human health is the polio virus. Polio caused paralysis and death in several thousand people a year until a vaccine was formulated in the 1950s. The genetic strand for the polio virus is composed of 7411 RNA bases. A single long poly-protein is formed as the RNA is translated within the cytoplasm of the human host cell. The poly-protein is eventually cleaved into ten proteins. Four of these form the capsid and six are involved in turning off the host’s own protein translation and in packaging new viruses for dispersal into the host. Only three of the four capsid proteins are visible from the outside of the icosahedron. The fourth protein is an interior protein.

The complex virus category includes viruses with icosahedron heads to which a helical tail is attached. Also within the complex category are a few viruses that change shape. They do not move from icosahedral to helical, rather they are strand-like structures that bend into numerous shapes. The Ebola virus falls into this category. The 900 nm-long virus can be straight, bent or folded. No matter the shape, Ebola causes so much internal hemorrhaging that 90% of its victims die. Two other complex viruses of note are the pox viruses and HIV. HIV has an enclosed cone shape while the poxes are brick-shaped.

Both the poliovirus and TMV are so-called naked viruses in which the capsid proteins are exposed to the environment. The lack of a lipid layer makes the naked viruses easier to identify and treat with anti-viral medications or vaccines. Many viruses, like the Ebola virus, HIV, and the influenza virus, are covered by a lipid envelope. The virus obtains this coating as it exits the host cell’s nuclear or cell membrane or wraps into the host’s endoplasmic reticulum. The envelope is like camouflage. The virus does alter the host’s membrane by adding a few of its own proteins, including a probe protein. This special protein aides the virus in locating a specific receptor on the host cell’s cell membrane. Herpes is an example of an enveloped icosahedron. Rabies is an enveloped helical virus. Since the envelope surrounding the capsid is partially composed of the host’s membrane, treating the virus by attacking the envelope would also attack the host’s cells. This feature makes enveloped viruses more difficult to treat or to vaccinate against.

Experiment Overview

Construct two scale-model viruses and determine the correct amount of model RNA to insert into each capsid.

Materials

Prelab Questions

1. One nucleotide of RNA or DNA is approximately 0.34 nanometers long. One millimeter equals 1,000,000 nm. If the genetic sequence for HIV is 9749 nucleotides, how long is the genome for HIV in nanometers? In millimeters?
2. A single HIV capsid is 120 nanometers (nm) in diameter. In order to make a model of HIV the capsid is scaled up to 120 mm. How long would a scaled model of the genome be? Hint: Refer to the Background section.

Safety Precautions

The materials used in this activity are considered nonhazardous. Please follow all normal classroom safety guidelines.

Procedure

Part A. Polio Virus

1. Cut out the icosahedral form on the Polio capsid sheet.
2. Make all folds by folding each solid line, ——, so the graphics remain on the outside of the shape.
3. Attach all but one of the adjacent cut lines to each other using tape.
4. The polio virus is 30 nm in diameter. Measure the diameter of the completed model capsid by inserting the ruler through the opening in the icosahedron. Record the diameter on the Virus Worksheet.
5. Using the information from the Background and Prelab Questions, calculate and record the length of the polio genome in nm.
6. Based on the diameter of the capsid model, calculate and record the length of the genome in the polio model.
7. Measure and cut a piece of string the correct length of the model genome.
8. Insert the model genome into the capsid model.

Part B. Tobacco Mosaic Virus

1. Wrap the 18" wire around a 100-mL graduated cylinder to create the helix shape (see Figure 2 in the Background section.)
2. Gently pull the looped wire off of the graduated cylinder.
3. Cut out the 21 model proteins on the TMV sheet.
4. Beginning about one inch from the end of the wire helix, wrap one model protein around the wire at the solid line (see Figure 4).
   {11107_Procedure_Figure_4}
5. Tape the tab to the underside of the protein model forming a protein “petal.”
6. epeat with the remaining proteins, placing each new protein next to the previous one.
7. Once all 21 proteins have been affixed to the wire use wire cutters to remove all but ½" of excess wire from the ends of the TMV capsid model.
8. Insert a ruler into the center of the TMV capsid model and adjust the helix to 60 mm tall.
9. The tobacco mosaic virus is 300 nm long. Using the information from the Prelab Questions, calculate and record the length of the TMV genome in nm.
10. The part of the capsid model made is 60 mm long, but only contains 21 proteins. A full-sized model would contain 2160 proteins and be a little over 6 meters long! Based on the length of the TMV capsid model, calculate and record the length of the model’s genome in nm.
11. Measure and cut a piece of string that is 100x smaller than your calculated length. This is the portion of genome that would be contained within this segment of the TMV model. Record the answer on the Virus Worksheet.
12. Loop the model genome within the TMV capsid and secure it at both ends of the TMV capsid with tape.

Student Worksheet PDF

11107_Student1.pdf