Upon completion of this module topic, you should:
- Be able to explain the importance of protein structure (primary, secondary, tertiary, quaternary) in relation to amino acid sequences and protein interactions with other proteins and small molecules.
- Be able to apply your understanding of protein structure, function, and interactions to the ways in which drugs (such as antibiotics) can interact with proteins and impact their functions and role in various physiological processes.
This is Part A, Protein Expression, under the module topic Protein Techniques. This topic part has two sections to review: Content Tutorial & Animations.
- DNA to Proteins (learn.genetics.utah.edu, HTML Page)
Once you are at the DNA to Protein web page, view the right side of the screen where it says “Proteins”. Click on the Tour the basics, What is a Protein? to review some general information about the role and production of proteins in the human body. When you have finished the Tour the basics, click on the Interactive Explore section below it to view the following animation, “Test Neurofibromin Activity in a Cell”. The animated activity allows you to see how normal cell division can be affected by a protein mutation. In the activity you will learn about the NF1 gene that codes for the protein neurofibromin and how it interacts with and regulates the function of the Ras protein that promotes cell division.
Overview of Protein Expression Systems: Workflow (Invitrogen)
The following flow chart has been designed by the Invitrogen company and provides a summary of the workflow involved in using a protein expression system to study and validate protein function and expression. Many of the techniques displayed in the flowchart for Sample Preparation, Protein Separation, Protein Characterization and Functional Validation will be explored in the protein module subsets b and c.
Reference: Invitrogen (www.invitrogen.com, HTML Page) “Protein expression enables analysis of protein structure and function. Use of recombinant proteins varies widely—from functional studies in vivo to large-scale production for structural studies and therapeutics. Using the best expression system for your protein and application is the key to your success. Solubility, functionality, speed, and yield are often the most important factors to consider when choosing an expression system. Invitrogen offers a wide variety of expression systems so you’ll be sure to find one that meets your needs. The following table highlights the characteristics of the most popular expression hosts. “ (Invitrogen) The second flowchart provided below was also designed by Invitrogen and organizes the various types of protein expression systems and vectors that can be used in vitro and in vivo to study protein function and expression. As you can see, the following hosts span a diverse array of organisms that include both prokaryotic and eukaryotic systems.
|Host Organism||Most Common Applications||Advantages||Potential Challenges|
Reference: Invitrogen (www.invitrogen.com, HTML Page)
1. Click on the following link to view the Howard Hughes Medical Institutes “Click and Learn” Biointeractive presentation, Visualizing Gene Expression (this presentation is also in the Nucleic Acids Module in the “Nucleic Acid Hybridization and Expression Analysis subset (III-c.).” The presentation includes important information about two important principles of detection for visualizing protein molecules, specificity and measurability. There is also subsequent material that includes methods for choosing a target by detecting proteins with antibodies and detecting messenger RNA with base pairing. In addition, there is information about methods for amplifying the detection signal and using reporter genes for transgenic technology.
- Visualizing Gene Expression (www.hhmi.org, Multimedia Page) Learn about the different ways scientists are able to detect when genes are being expressed in various tissues.
2. The following Animation provides a comparison between prokaryotic and eukaryotic organisms for the processing of gene information. The animation complements the comparison flowchart above from the Invitrogen website.
- McGraw Hill Animation (highered.mcgraw-hill.com, Multimedia Page)
When you access the website, click on the animation “Processing of Gene Information – Prokaryotes versus Eukaryotes”.
Proteins & Proteomics
This is Part B, Proteins & Proteomics, under the module topic, Protein Techniques. This topic part has three sections: Content Tutorial, Animationsand Activities.
Click on the following link to view the following Annenberg Media, Rediscovering Biology online textbook, content tutorials on proteins and proteomics. Use the arrows on the bottom of the webpage to navigate through the content webpages. The links provided below will allow you to view relevant selections from the online textbook.
- What is Proteomics? key terms: proteome, alternative splicing (www.learner.org, HTML Page)
- Introduction to Protein Structure key terms: primary structure, secondary structure, tertiary structure, quaternary structure, domains, domain shuffling, motifs (www.learner.org, HTML Page)
- Determining Protein Structure key terms: x-ray crystallography, nuclear magnetic resonance (www.learner.org, HTML Page)
- Structure and Function Relationships of Proteins key terms: active site, ligand (www.learner.org, HTML Page)
- Genomics-Based Predictions of Cellular Proteins key terms: glycosylation, phosphorylation, protein sorting, kinases, phosphatases (www.learner.org, HTML Page)
- Proteomics & Drug Discovery key terms:virtual ligand screening (www.learner.org, HTML Page)
- Identifying Protein Interactions key terms: interaction domains, catalytic domains (www.learner.org, HTML Page) (Reference: Annenberg Media, Rediscovering Biology)
1. Annenberg Media, Rediscovering Biology: Proteins & Proteomics
View the following animations to see depictions relating to protein interactions, structure and function. (Reference: Annenberg Media, Rediscovering Biology)
- The Evolution of Protein A depiction of how evolution can affect how proteins interact with other proteins.
View Quicktime Movie (www.learner.org, Multimedia Page)
- The Three-Dimensional Structure of a Protein A depiction of the subsets of a protein structure.
View Quicktime Movie (www.learner.org, Multimedia Page)
- Virtual Ligand Screening in Drug Design Shows how a computer program can be used to fit potential drug molecules into a site of interest on a protein.
View Quicktime Movie (www.learner.org, Multimedia Page)
Brief descriptions of animations: a. The Evolution of Protein-Protein Interactions A depiction of how evolution can affect the way proteins interact with other proteins. b. The Three-Dimensional Structure of a Protein A depiction of the subsets of a protein structure (primary, secondary, tertiary, quaternary) c. Virtual Ligand Screening in Drug Design A depiction of how a computer program can be used to fit potential drug molecules into a site of interest on a protein.
2. Howard Hughes Medical Institute Biointeractive Animations: Protein Composition, Structure, Function, Interactions
a. Click on the following link to view an animation about protein structure and protein structure modulations that can result from protein interactions with other proteins and molecules.
- Using Small Molecules to Modulate a Protein
This animation illustrates how a small molecule binds to a protein. As a result of the binding, the protein alters its shape and becomes inactivated. 1 minute 10 seconds Large: Play: MOV / WMV Small: Play: MOV / WMV
b. This second animation focuses on the importance of protein structure and the devastating effects that can result from changes in protein structure particularly for the protein hemoglobin. In the specific case of sickle cell anemia, the disease results from a gene mutation that causes an amino acid substitution in the protein hemoglobin. The amino acid change affects the structure of hemoglobin and results in a phenotypic change in the shape of red blood cells.
- Sickle cell anemia
Sickle cell anemia is a genetic disease that affects hemoglobin. A single nucleotide change in the hemoglobin gene causes an amino acid substitution in the hemoglobin protein from glutamic acid to valine. The resulting proteins stick together to form long fibers and distort the shape of the red blood cells. 59 seconds Large: Play: MOV / WMV Small: Play: MOV / WMV
c. The third animation applies to protein interactions and the ways in which drugs such as antibiotics can interact with proteins and impact protein functions and the role(s) that particular proteins play in human physiological processes.
Rapamycin is a small molecule originally isolated from nature. It has antibiotic and immunosuppressive properties. It also allows two proteins which do not normally interact to bind together in the cell, which causes problems in the nutrient-sensing pathway. 1 minute 9 seconds Large: Play: MOV / WMV Small: Play: MOV / WMV
d. Click on the following link to view the Howard Hughes Medical Institute’s biointeractive tutorial about the molecular structure and function of a very large protein known as a proteasome.
- Anatomy of a Proteasome (hhmi.org, Multimedia Page) e. Click on the following link to view the biointeractive tutorial about a very important cancer gene, p53 known as “The Guardian of the Genome”. p53 (hhmi.org, Multimedia Page)
CASE STUDY – Proteomics & Drug Discovery
Reference: Annenberg Media, Rediscovering Biology “Designing Cancer Drugs”
a. Before you begin the case study, read the following article from the National Cancer Institute (U.S. National Institutes of Health: www.cancer.gov), “FDA Approves Important New Leukemia Drug: Offers Further Proof of Principle for Molecular Targeting in Cancer Treatment”.
- Gleeve Press Release (www.cancer.gov, Multimedia Page)
Reference (www.cancer.org, Multimedia Page)
b. Click on the following link to view the animation to see how the drug Gleevec mimics ATP to bind a cancer-causing enzyme in order to block its function.
Gleevec (1 minute 3 seconds)
Gleevec mimics ATP, binds to a cancer-causing enzyme and blocks its function. Large: Play: MOV / WMV Small: Play: MOV / WMV | Learn More c. Now complete the Case Study: Annenberg Media Rediscovering Biology To begin, click on the following link to view the case study homepage. As you will see, there are related materials (tutorials and videos) provided as links that are recommended for viewing prior to beginning the activity. When you have finished viewing the related materials begin by clicking on the green “Go” arrow below to “Launch Case Study”. The case study will provide you with an introduction and overview to the activity. The specific steps involved in the case study are listed below.
Case Study Steps (www.learner.org, Multimedia Page) 1. Focus on a disease 2. Identify the cause of a disease 3. Identify a drug target 4. Screen to find a lead compound 5. Modify and retest the lead compound 6. Lab and animal testing 7. Human clinical trials (Phases I, II and III) 8. New drug approval 9. Post-marketing surveillance (Phase IV)
Launch Case Study (www.learner.org, Multimedia Page) ““Designing drugs” sounds trendy, but it accurately describes how some remarkably effective drugs are developed. In this case study we’ll follow the steps of drug design, from initial research on the targeted disease, to the drug’s use in humans. Gleevec (STI-571) is our example. We’ll see how drug design exploits the structure of a protein that causes a disease. We’ll consider the qualities of an effective drug, like high specificity, low toxicity, and ease of delivery. Animations will show how translocation creates the oncogene that causes chronic myelogenous leukemia, and how Gleevec inhibits the enzyme encoded by the oncogene.” (Reference: Annenberg Media, Rediscovering Biology)