ACTIVITY 4

PROTEIN MALFUNCTION AND DISEASE: THE EXAMPLE OF SICKLE CELL ANEMIA

Activity Overview:

Key concepts:
Mutations in DNA result in changes in the sequence of amino acids of a protein (its primary structure). Thus, mutations may lead to changes in the way a protein functions and can become the molecular cause of illness.

Students continue their exploration of the DNA coding of protein sequence. They use the Molecular Workbench model Mutations; Substitutions and Deletions to compare the effect of two different types of mutations in DNA, one caused by a nucleotide substitution and another caused by a nucleotide deletion. Students bring what they have learned in their investigation of the coding of proteins and the relationship between protein's structure and function to the case of Sickle Cell Anemia. They reproduce a critical piece of hemoglobin structure using the Molecular Workbench: Modeling Hemoglobin model to explore changes in folding much like the change in hemoglobin in a patient with Sickle Cell disease. Finally, they take a post-test.

Learning Objectives:

Students will be able to:

Compare the effect of nucleotide substitutions and deletions on protein structure;
Reason about the molecular origin of disease;
Relate the change in the structure of proteins to changes in their function and possible implications for human health.

Macro to Micro Connection:

Students will relate some macroscopic symptoms characteristic of human disease to microscopic changes in the primary structure of nucleic acids and proteins.

Conceptual Prologue

Mutations and Illness: Mutations can result from various kinds of damage to the structure of DNA. Substituting one nucleotide for another often makes no significant change in the shape of a protein, unless it occurs at a critical location. Due to the redundancy of the genetic code, in which several different codons in fact "mean" the same amino acid, many changes have no effect. Some substitutions, on the other hand, lead to change in the genetic code that brings about replacement of an hydrophilic amino acid to a hydrophobic one or vice versa. This can have a great effect.

When a deleted or substituted amino acid is located in a critical position in the protein, it will affect the shape of the protein. That in turn can affect the way the protein works.

Deleting a nucleotide, on the other hand, is likely to change many codons located immediately after the deletion. As the codons are read sequentially by the protein assembly line, all of the codons after the deleted nucleotide that are changed will generate the placement of "wrong" amino acids in the protein chain. Such a change typically leads to the appearance of a non-functional protein; if it was essential for the living cell protein such deletion would be a lethal mutation.

A change in sequence change in shape change in activity.

Many medical conditions reflect a disorder of a protein's function, e.g. Sickle Cell Anemia, PKU, Hemophilia, Night Blindness, B1 Deficiency. Such illnesses are the result either from flaws of metabolism or from damages of cellular structures, and they often originate in mutations in DNA.

Sources of Mutation Mutations can either be inherited or acquired during our lifetime. Some kinds of cancer may be a combination of an inherited mutation and a mutation that we develop during our lifetime. Changes in proteins that result in disease can be the result of inborn genetic errors, so that for a certain person, DNA in every cell will always carry the mutation for the wrong protein. Changes in DNA can also result from the exposure of an organisms or any group of cells to environmental factors, such as UV rays of ionizing radiation, such as of the radiation in cosmic rays or nuclear reactions. These mutations can have different consequences for an organism, depending in what cells and tissues they occur. Mutations that occur in sperm cells or eggs can be passed to the next generation.

Sickle Cell Anemia Some people inherit a specific substitution of a nucleotide in the gene that controls the sequence of amino acids in hemoglobin. These people inherit Sickle Cell Anemia. The hemoglobin molecules in people with this disease are folded in such a way that they stick to each other, making long fibers that damage red blood cells.

The critical event - the formation of the fibers - occurs because the mutation replaces glutamic acid in the 6th position of the protein chain with valine, an amino acid with different properties. This replacement changes the shape of the protein: a small protrusion appears on the surface of one of the proteins making hemoglobin. This bump fits exactly into the existing "pocket" on the surface of the adjacent protein. The two proteins "clump" together, then the third clumps...This creates a kind of domino effect leading to the formation of long fibers made of many millions of damaged hemoglobin molecules.

Activity Design and Execution

Major Science Concepts	Mutation, insertion, deletion, substitution, genetic disease, DNA to Protein structure
Assumed Previous Knowledge:	Amino acids, charge, codon, DNA
Time:	1-2 50 minute classes
Technical Requirements:	There are two models in this activity, and they can be launched in one of two ways: 1. From your browser. Click the link below.: * Molecular Workbench: Mutations [http://xeon.concord.org:8080/modeler/webstart/protein/mutations.jnlp] * Molecular Workbench: Hemoglobin [http://xeon.concord.org:8080/modeler/webstart/protein/hemoglobin.jnlp] 2. By going through the Molecular Workbench application on your computer (workbench.jar), click the following links: Student Pages, Protein Folding, Mutations or Hemoglobin. It may take a short while to launch the Molecular Workbench the first time.
Supporting Materials:	* Background: Molecular Machines and Tools Chart [PDF version] * Worksheet: Mutation (Student) [PDF version] * Worksheet: Mutation (Teacher) * Worksheet: Protein Malfunction and Disease: Making a Sickle Cell Mutation (Student) [PDF version] * Protein Malfunction and Disease: Making a Sickle Cell Mutation (Teacher) Post Test Post Test Answers
Advanced preparation	Print or bookmark support materials Prepare model for access (See above) Visit and bookmark Student Index

Investigative question: How can changing one nucleotide cause a disease?

Steps

1. Hemoglobin: A Molecular Machine. Discuss the Molecular Machines and Tools Chart, considering hemoglobin as a molecular machine, together with the many other critical and specific jobs proteins perform in the cell. Ask students to think about what change in the body would occur if any of these molecular machines on the chart would be mutated. What might happen if those molecules could not perform their job well or at all? (Accuracy here is less important than their attempt to think through some implications. With a deformed hemoglobin, a cell could not carry oxygen where it is needed.) You might also discuss some environmental sources of mutation, e.g. radiation.

2. Exploring two ways mutations occur: substitution and deletion. Distribute the Mutation Worksheet (Student). In this activity students model DNA mutations and look at the affects on the shape of the protein fragment. It is important to discuss with the students that the model does not show the intermediate steps of translation and transcription. Instead the model shows the cause and effect between a mutation and protein structure. Nonetheless, students will be expected to analyze what is happening keeping the intermediary steps in mind. Make sure that you discuss student responses after they have completed the activity.

3. Understanding Sickle Cell Anemia. This concludes students exploration into the molecular mechanism related to Sickle Cell Disease. Students compare the DNA code for normal and sickle cell hemoglobin, observe the difference in protein structure and learn about how the difference in structure is related to the presence of disease symptoms. Distribute the worksheet Malfunction and disease: Making a Sickle Cell Mutation.

You might want to discuss why genetic therapy might be a promising method for helping with this disease, or have students research other treatments.

4. Finally, distribute the Post-test.

Extensions

Explore the Visual Story of Hemoglobin.

Have students read about PKU disease. Discuss what the disease has in common with Sickle Cell Anemia. (It is inherited, derived from a single point mutation in the DNA.) Then consider ways in which the disease differs from Sickle Cell Anemia. (A critical liver enzyme is malfunctioning in PKU disease; in Sickle Cell Anemia, the hemoglobin is misshapen, aggregating in long fibers damaging the red blood cell.) http://www.concord.org/~barbara/homs/pku2.html

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