The 2017-2018 Theme:

Influenza Virus Proteins Involved in
Getting the Virus Into and Out of Host Cells

Be extra sure to explore the information in all three tabs below


Influenza Protein Structure The 2017-2018 Proteins

Welcome!
To the 2017-2018 Science Olympiad Protein Modeling Event Webspace

This year’s event focuses on two proteins found on the surface of the influenza virus. One protein, hemagglutinin, undergoes a dramatic pH-induced conformational change that allows the viral genetic material to enter the cell. The other protein, neuraminidase, is important in helping the virus to both 1) get through the sticky mucus layer to enter the cell and 2) allow the replicated viral particles to escape the cell. Neuraminidase is the target for antiviral agents such as Tamiflu.

We’ve provided a LOT of resources to help you to understand the proteins in the context of the virus life cycle. As you work through the materials, you will discover some questions in italics. These questions will provide you the opportunity to explore some topics in greater depth....Sometimes there will be specific answers that you can easily find on the Internet, but other questions will require some thought and/or analysis of information you already have. Science is all about asking questions – and you will probably come up with some great questions on your own that you’ll also want to explore. But the questions we’ve included will help to guide you into thinking like a scientist. So – happy exploring!



A Word On Terminology

As you probably know, studying science is like learning another language! Some scientific terms are only used in science, and other terms that you MAY be familiar with in everyday life have a DIFFERENT meaning when used in science. We’ve created some videos to introduce some of the terms you’ll encounter as you explore proteins found in the influenza virus. Click on each link below to expand the video:

Genetic Drift and Genetic Shift
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Genetic Shift - Production of new virus due to mixing of genomes of two or more viruses.

Genetic Drift - Minor changes in viral DNA resulting in small changes in protein structure.

Virulence
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Virulence - The ability of the microbe to damage the host. (How sick will the virus make you?)

Transmissibility
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Transmissibility - Measure of how easily the virus can be passed from individual to individual within a population.

Host Susceptibility
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Host Susceptibility - how likely a host is to become ill if exposed to a microbe. Often depends on other factors, such as age or whether there are underlying medical conditions..

Herd Immunity
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Herd Immunity - When a large portion of the population is resistant to a microbe, the microbe is not able to cause an epidemic.

Sialic Acid Linkages
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Sialic Acid Linkages - Sialic acid sugars are attached to galactose on either the 3rd carbon or the 6th carbon of the galactose, which is attached to the 2nd carbon of the sialic acid. Hemagglutinin molecules recognize one of these two linkages.

Heptad Repeat
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Heptad Repeat - Repeating pattern of seven amino acids that suggests that this portion of a protein forms an alpha helix that joins with other alpha helices to create a coiled coil.



Introduction

Perhaps you’ve experienced a conversation similar to this:

We often use the term “flu” when we talk about short-term illness – either respiratory or digestive - that has us “under the weather”, but influenza (the flu) is actually a fairly common respiratory infection that can be quite serious – and in some cases fatal. If you’ve ever been unfortunate enough to be infected with the influenza virus, you will recall the achy muscles that make you feel like you've been run over by a train.

We’ll explore this virus in more detail, including its structure, how it gets into and out of the host cell, and why health care officials are concerned about newly developing viruses. Influenza virus has been widely studied by scientists for many years. As our knowledge of other viruses grows, we’ve discovered that many viruses use the same mechanisms to infect cells. So once you know about influenza virus, you’ll be able to apply what you know to other viruses, including Ebola, Zika and HIV.



History of Influenza Pandemics


Policemen wearing masks provided by the American Red Cross in Seattle, 1918. From: Illinois Pandemic Flu Organization

The flu is a common human ailment throughout the world. Each year in the United States alone, 5-20% of the population will be infected with the flu. Of these, more than 200,000 people will be hospitalized due to flu related complications such as high fever, pneumonia, ear and sinus infections and dehydration. Approximately 36,000 people will die of such complications.

Over the past 100 years, four separate flu pandemics have shaped our understanding of this infectious disease: the 1918 Spanish Flu, the 1957 Asian Flu, the 1968 Hong Kong Flu and the 2009 Novel H1N1 Flu. Explore the links below to learn more about these pandemics.



1918 Spanish Flu
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Policemen wearing masks provided by the American Red Cross in Seattle, 1918. From: National Museum of Health and Medicine, Armed Forces Institute

The 1918 flu pandemic was the worst recorded in global history. Approximately 500 million people were infected with the virus, killing an estimated 50 million people worldwide. The initial outbreak of this virus occurred in the spring of 1918, causing an average number of illnesses and deaths. When the virus “returned” for the fall flu season, it had mutated and become more virulent.

Perhaps the most disturbing trait of this virus was not the efficiency with which it infected and killed its victims, but the groups that it attacked. Those born before 1889 appeared to have a partial immunity to the disease and were not as severely affected, but the very young and those aged 20-40 years suffered the greatest casualties.

Descendants of this virus strain still appear in seasonal influenza outbreaks, but do not show the same virulence. Research has suggested that the combination of the 1918 hemagglutinin, neuraminidase and polymerase was such that the virus was capable of replicating much faster, contributing to its increased virulence (Shen et al., 2009; Tumpey, 2005).

References

Shen, J., Ma, J., and Wang, Q. (2009). Evolutionary Trends of A(H1N1) Influenza Virus Hemagglutinin Since 1918. PLoS ONE 4, e7789.

Tumpey, T.M. (2005). Characterization of the Reconstructed 1918 Spanish Influenza Pandemic Virus. Science 310, 77–80.

1957 H2N2 Asian Flu
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Dr. Joseph Ballinger gives Marjorie Hill, a nurse at Montefiore Hospital, the first “Asian flu” vaccine shot to be administered in New York, on Aug. 16, 1957. The vaccine was judged 45 to 60 percent effective. From: The Seattle Times

The 1957 Asian Flu outbreak, while not as deadly as the 1918 Spanish Flu (it is estimated to have claimed 2 million lives around the world), was significant in that it was the first known occurrence of the H2N2 influenza variant. The strain originated when an avian influenza strain recombined with a preexisting human influenza strain. The virus was first detected in Guizhou, China in 1956. By 1957 a vaccine had been created against the strain, stemming its further spread. (The very first flu vaccines were developed in 1938.)

1968 H3N2 Hong Kong Flu
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Chinese Red Guard members were ordered by Chairman Mao to cover their mouths to protect against flu germs in 1968. From: Signs of the Times

The 1968 Hong Kong Flu was the first recorded appearance of the H3N2 influenza A subtype. It was a descendant of the 1957 H2N2 Asian Flu strain by means of antigenic shift, or a recombination of multiple influenza strains in a single host. In the case of H3N2, it seems likely that strains of human and avian influenza were recombined in a swine host. Descendants of this strain evolved into the annually circulating seasonal flu strains.

2009 Novel H1N1 Flu
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Electron micrograph of influenza virus. From: Revolution Health

Students at a University of London class in Mexico City wear masks to protect them against novel H1N1 flu in 2009. From: National Public Radio

First detected in the spring of 2009 in Mexico and the United States, Novel H1N1 Flu was a new recombinant virus containing genes from human, avian and swine influenza strains. Although similar to the 1918 flu strain, the novel H1N1 flu is not as virulent, and it is sensitive to antiviral drugs. This strain continues to circulate widely as a seasonal flu strain, but because many people get annual flu vaccines, they have developed an immunity to this particular strain.


For more information about the history and current status of the H1N1 flu, check out:



Current Status of Influenza Strains

As influenza is a continually evolving and seemingly ever present virus, our knowledge, understanding and research regarding this disease are also continuously growing. Explore below for more information about the current status of each of the listed flu strains.

Seasonal Influenza
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Model of influenza virus built with 3D printing technologies. From: 3D Molecular Designs

Seasonal flu differs from pandemic flu in several ways: Seasonal flu outbreaks occur annually, following predictable patterns. This is due to antigenic shift – minor mutations in existing flu strains that allow the virus to evade the host’s immune system. The mutations typically occur in the gene that codes for the hemagglutinin protein, resulting in subtle changes in the protein structure. As these changes are slight, the host immune system can still recognize the virus as similar to previously circulating strains.

Because of this continued exposure to nearly identical virus strains, the general population is capable of building up some immunity, called “herd immunity”. This means that healthy adults are not generally at risk for any serious complications, though the very young, the elderly and those with additional health complications may suffer more severely from seasonal flu infection. It is possible (though tricky) to predict which strains of flu will be circulating during a given season, and vaccines are created to help prevent and limit the spread of the virus.

Pandemic flu occurs rarely, unpredictably, and may cause complications even in healthy individuals. It is also unlikely that a vaccine could be developed for the early stages of the pandemic, and there is a chance that antivirals would be less effective in treating infection.


The following websites provide additional information about seasonal influenza:

Avian Influenza Viruses
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Wild waterfowl are the natural hosts of influenza viruses. Avian viruses bind to and infect the digestive tract of birds, and viruses are transmitted through direct contact with fecal matter on contaminated ground, food or water. Birds infected with the influenza virus typically do not get sick.

The H5N1 virus, unlike most avian influenza viruses, is highly pathogenic, typically leading to death of most birds within 48 hours. This pathogenicity is due to a specific variation in the hemagglutinin protein. As you learn more about influenza viruses, you might want to identify what variation in the hemagglutinin protein leads to highly pathogenic strains of the influenza virus.

In the past 20 years, persons working in close proximity to infected birds have become infected with avian strains of the influenza virus. This is unusual because although avian and human influenza viruses both recognize sialic acid sugars on the surface of cells, the way the sialic acids are attached to the sugar chain differs enough between humans and birds that the avian virus can’t readily bind to human cells. Just changing a couple of amino acids in the hemagglutinin protein can determine whether the virus binds better to human or bird receptors (Connor et al., 1994), so healthcare providers and scientists have been closely tracking human cases of avian flu to see if and when these viruses are transmitted directly from one human to another. To date, any human-to-human transmission has occurred only rarely. This change in the behavior of the virus could signal the beginning of a new pandemic for which humans are not immunologically prepared.

Researchers are working to develop a vaccine against H5N1 avian virus. Although there are a number of antiviral drugs that target the neuraminidase or M2 viral proteins, many influenza strains have become resistant to these drugs, and the fear is that avian strains will also develop resistance to these drugs. The rapid mutation rate in influenza viruses is partly due to the fact that they are single-stranded RNA viruses and partly due to the error-prone nature of the viral RNA polymerase.

Reference

Connor, R.J., Kawaoka, Y., Webster, R.G., and Paulson, J.C. (1994). Receptor specificity in human, avian, and equine H2 and H3 influenza virus isolates. Virology 205, 17–23.


The following websites provide additional information about avian influenza:



"Life" Cycle of a Virus

Viruses come in many shapes and sizes! All viruses have genetic material – either DNA or RNA (but not both). The genetic material can be single-stranded or double stranded. This genetic material is packaged in a protein coat called a capsid. Some capsids consist of multiple copies of a single protein; others are composed of a variety of different proteins. Some viruses pick up a membrane coat as they leave the host cell. This membrane is part of the host cell, but viral proteins can be expressed on the surface. Viruses that have an outer membrane are called enveloped viruses, and the membrane is called the envelope.



Check out this video that describes how viruses enter and leave cells.



Overall Viral Structure

Click on the buttons below for a definition of each part of the influenza virus capside, as well as to see it marked on the image to the right.

Viral Envelope
M1 Matrix Protein
Hemagglutinin
Neuraminidase
RNA Packing Protein
Viral RNA
M2 Channel


This video uses a 3D model to introduce you to the influenza virus:



Check out Vincent Racaniello’s Virus Blog about the structure of the influenza virus link:
http://www.virology.ws/2009/04/30/structure-of-influenza-virus/



This video uses 3D models to introduce you to the hemagglutinin protein:



Check out the Protein Databank's Molecule of the Month on hemagglutinin:
http://pdb101.rcsb.org/motm/76


Check out these videos by Dr. Stephen Harrison at Harvard Children's Hospital talking about the HA protein structure and fusion:

An interesting article:


This video uses 3D models to introduce you to the neuraminidase protein:



Check out the Protein Databank's Molecule of the Month on Neuraminidase:
http://pdb101.rcsb.org/motm/113



Vaccines against Influenza

A vaccine is a dead or inactivated organism (or products derived from them) that are introduced into the body to “train” the immune system to recognize the virus. While these particles do not illicit an illness, the immune system still recognizes them as invaders. Once identified, the immune system creates proteins known as antibodies that circulate in the blood. The antibodies recognize and bind to the hemagglutinin protein exposed on the surface of the virus and prevent the virus from infecting cells. Once the immune system learns to recognize a specific influenza strain, it maintains “memory cells” that can quickly mount an immune response if the body is exposed to that virus again.

Each year, the Centers for Disease Control (CDC) and the World Health Organization (WHO) analyze which influenza strains are circulating in the population and predict strains that are likely to be circulating in the coming flu season. Typically, three strains of influenza virus are chosen to be included in the seasonal flu vaccine: one influenza A (H3N2) virus, one influenza A (H1N1) virus, and one influenza B virus. When these have been selected, the production of vaccines can begin.

What influenza strains were recommended for the current season (northern hemisphere winter)?



How Vaccines are Made

Because viruses can’t replicate on their own, the virus is amplified by injecting virus into the amniotic fluid of an embryonated chicken egg (chicken embryo) where it multiples for several days. Viruses from this egg are injected into additional eggs, repeating the process in millions of eggs to produce enough virus to create the vaccine. The amplification process takes approximately six months and requires multiple quality control checks to ensure that the virus does not mutate during production. Once the virus has been amplified, the fluid containing the viruses is removed from the eggs and multiple purification steps concentrate the virus and remove any proteins from the chicken. The virus is then treated chemically to inactivate it so that it can’t cause infection. The three purified, inactivated viruses are then mixed in a solution that can be injected. The vaccine must be approved by the FDA before it can be administered.



Check out these relevant Molecule of the Month topics:

What is the advantage of broadly neutralizing antibodies in preventing influenza infections?

In order to truly appreciate and successfully model this year's Protein Modeling Event structures, a thorough understanding of protein structure is needed. This section will explore these amazing macromolecules in more detail using suggested physical model kits, online resources and additional websites.

Protein Structure

Proteins are long linear sequences of amino acids
that fold into complex 3-dimensional shapes
following basic principles of chemsitry.

In this first of two videos on protein folding, Tim Herman, Ph.D. from the MSOE Center for BioMolecular Modeling uses the Water Cup from 3D Molecular Designs to demonstrate some basic principles of chemistry.




In this second of two videos on protein folding, Tim Herman, Ph.D. from the MSOE Center for BioMolecular Modeling uses the Amino Acid Starter Kit from 3D Molecular Designs to demonstrate how the basic principles of chemistry directly affect protein folding.




In this video, Tim Herman, Ph.D. from the MSOE Center for BioMolecular Modeling uses the Alpha-helix Beta-sheet Construction Kit from 3D Molecular Designs to demonstrate the form and function of secondary structures in proteins.






Protein Databank (www.rcsb.org) Resources

Learn about protein structure and function with this overview printout and video developed by the RCSB Protein Databank.

Protein Structure Jmol Tutorials

The Protein Structure Jmol Tutorials walk through the four levels of protein structure using interactive Jmol molecular visualizations, including real protein examples with interactive controls.

3D Molecular Designs Educational Kits

These engaging, hands-on kits make learning protein structure basics easy. Users will fold a protein while exploring how the chemical properties of amino acids determine its final structure.


Click to Open the Final Tab: This Year's Proteins

Rule Changes!

If you’ve participated in the Science Olympiad Protein Modeling Event in previous years, please note that there are a few big changes to the rules. Refer to the official rules for explicit details, but the changes include:

  1. size limitations for pre-build models
  2. size and format of card detailing creative additions to the pre-build model
  3. requirement that models be sturdy enough that judges can pick up and rotate models for judging
  4. no pages, printed or hand written, can be brought into the onsite competition


Here’s a video that provides a little explanation of the rule changes:



Here is a sample notecard describing creative additions (modeling beta globin):



The table below lists the proteins that will be featured as the 2017 pre-build as well as the various on-site builds at invitational, regional, and state competitions.

Be Sure to Click on Each Protein Listed Below to reveal additional resources that you should explore as you prepare for each level of competition. In addition to providing the pdb file for the protein that will be featured, these links also provide a copy of the original research paper (the "primary citation") that reported the protein's structure. Questions regarding the paper and the structure presented in the PDB file will be asked on the exam you will complete at each competition.

Pre-Build Model Influenza Hemagglutinin 1htm.pdb
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Coordinates for the Model

The 2017-2018 Pre-Build Model should represent amino acids 40-153 of chain D of the influenza hemagglutinin protein based on the PDB file 1htm.pdb.

You can access the Pre-Build online design environment at http://cbm.msoe.edu/scienceOlympiad/designEnvironment/prebuild.html. Also study what types of additional features could be highlighted in the pre-build model in Section 3.

Background Information

A copy of the “primary citation” (the original paper that reported this structure) is provided below. You are not expected to read and understand the entire paper. However, reading the introduction and discussion and reviewing the figures in the paper should provide a good introduction to this protein. This structure shows the shape of the protein after membrane fusion. Choose to display features that tell a story about how the structure relates to the function of the protein.

Bullough, Per A., Hughson, Frederick M., Skehel, John J. and Wiley, Don C. 1994. Structure of the influenza haemagglutinin at the pH of membrane fusion. Nature 371:37-43.

Paper Abstract:

Low pH induces a conformational change in the influenza virus haemagglutinin, which then mediates fusion of the viral and host cell membranes. The three-dimensional structure of a fragment of the haemagglutinin in this conformation reveals a major refolding of the secondary and tertiary structure of the molecule. The apolar fusion peptide moves at least 100 Å to one tip of the molecule. At the other end a helical segment unfolds, a subdomain relocates reversing the chain direction, and part of the structure becomes disordered.

Additional Resources

This News and Views article provides insight into how hemagglutinin functions, based on this structure as well as the hemagglutinin structure used to create your pre-build model.

Stuart, David. 1994. News and Views: Docking mission accomplished. Nature 371:19-20.

Check out the Protein Databank's Molecule of the Month on hemagglutinin:
http://pdb101.rcsb.org/motm/76

Check out these videos by Dr. Stephen Harrison at Harvard Children's Hospital talking about the HA protein structure and fusion:

Regional On-site Model Influenza Hemagglutinin, Pre-fusion 5hmg.pdb
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Coordinates for the Model

The 2017-2018 Regional On-Site Model will be taken from influenza hemagglutinin based on the PDB file 5hmg.pdb.

Background Information

A copy of the “primary citation” (the original paper that reported this structure) is provided below. You are not expected to read and understand the entire paper. However, reading the introduction and reviewing the figures in the paper should provide a good introduction to this protein.

Wilson, I.A., Skehel, J.J. and Wiley, D.C. 1981. Structure of the haemagglutinin membrane glycoprotein of influenza virus at 3 Å resolution. Nature 289:366-373.

Paper Abstract:

The haemagglutinin glycoprotein of influenza virus is a trimer comprising two structurally distinct regions: a triple-stranded coiled-coil of α-helices extends 76 Å from the membrane and a globular region of antiparallel β-sheet, which contains the receptor binding site and the variable antigenic determinants, is positioned on top of this stem. Each subunit has an unusual loop-like topology, starting at the membrane, extending 135 Å distally and folding back to enter the membrane.


Check out the Protein Databank's Molecule of the Month on hemagglutinin:
http://pdb101.rcsb.org/motm/76

Check out these videos by Dr. Stephen Harrison at Harvard Children's Hospital talking about the HA protein structure and fusion:

State On-site Model Influenza Neuraminidase 2hu4.pdb
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Coordinates for the Model

The 2017-2018 State On-Site Model will be taken from influenza neuraminidase based on the PDB file 2hu4.pdb.

Background Information

A copy of the “primary citation” (the original paper that reported this structure) is provided below. You are not expected to read and understand the entire paper. However, reading the introduction and reviewing the figures in the paper should provide a good introduction to this protein.

Russell, Rupert J., Haire, Lesley F., Stevens, David J., Collins, Patrick J., Lin, Yi Pu, Blackburn, G. Michael, Hay, Alan J., Gamblin, Steven J. Gamblin and Skehel, John J. 2006. The structure of H5N1 avian influenza neuraminidase suggests new opportunities for drug design. Nature 443:45-49.

Paper Abstract:

The worldwide spread of H5N1 avian influenza has raised concerns that this virus might acquire the ability to pass readily among humans and cause a pandemic. Two anti-influenza drugs currently being used to treat infected patients are oseltamivir (Tamiflu) and zanamivir (Relenza), both of which target the neuraminidase enzyme of the virus. Reports of the emergence of drug resistance make the development of new anti-influenza molecules a priority. Neuraminidases from influenza type A viruses form two genetically distinct groups: group-1 contains the N1 neuraminidase of the H5N1 avian virus and group-2 contains the N2 and N9 enzymes used for the structure-based design of current drugs. Here we show by X-ray crystallography that these two groups are structurally distinct. Group-1 neuraminidases contain a cavity adjacent to their active sites that closes on ligand binding. Our analysis suggests that it may be possible to exploit the size and location of the group-1 cavity to develop new anti-influenza drugs.

Check out the Protein Databank's Molecule of the Month on Neuraminidase:
http://pdb101.rcsb.org/motm/113



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