Authors

Margaret Waterman, Southeast Missouri State University

William Coleman, University of Hartford

Robert Barber, University of Wisconsin - Parkside

Kathleen Duncan, Foothill College

Rick Cowlishaw, Southwestern College

Christopher Harendza, Montgomery County Community College

Abstract

6th Member: Chris Smith, Univ Calif, San Diego 7th Member; Marlene Kayne, The College of New Jersey Abstract Applications of bioinformatics to upper level classes are in their infancy and this approach is even less apparent in the introductory level courses. We propose the implementation of specific exercises that will allow students to utilize genomic databases to investigate simple examples of the linkage between gene sequence, protein structure, and the resulting normal and abnormal function. A tractable example for non-majors would be the gene and protein for rhodopsin and for majors the gene and protein tritin (and related proteins), which are antifungal agents, in grains. Goals of Integration of Bioinformatics tools: 1. Develop exercises from the perspective of a non-major that utilize genomics and proteomics as a tool for discovering and exploring biological concepts. For modules that use human disease as the hook for student interest, include assessment of the molecular basis for inherited disease using OMIM as a platform. 2. Develop exercises from the perspective of introductory biology students that utilize genomics and proteomics discovering and exploring biological concepts. Enhance current wet laboratory exercises with bioinformatics analyses that provide a deeper understanding of the molecular basis of biological phenomenon. Non-majors example: These lab modules would lead students through different levels of understanding biological concepts. Begin with topics familiar to non-majors; for example, the structure of the human eye, genetic diseases (cystic fibrosis, muscular dystrophy, sickle cell anemia) or current event topics (SARS, HIV, chronic wasting disease in deer). After introducing a familiar topic allow students to discover connections between DNA sequence, protein structure, protein function and gross phenotype. Next, use bioinformatics tools to allow students to discover how molecules have changed over time and what kind of similarities exist between diverse organisms (i.e., evolution and phylogeny). On 6/5/2003 a file was created that outlines some of the issues regarding the nonmajors component of the project. I could not upload the file directly, and so you will have to view it at http://faculty.mc3.edu/charendz/charend.htm You will need the authorware web player plugin to see it, if you do not already have it loaded. It can be obtained at If you do not already have it, you will need the Macromedia Authorware Webplayer to see this http://www.macromedia.com/software/authorware/productinfo/webplayer/ This part submitted by Chris Harendza Majors example: These lab modules approach the same topics but at a deeper level asking students to make connections between traditional wet labs and newer bioinformatics analysis. Students first explore the molecular tools used to purify and study molecules. Next students explore the connection between structure and function by manipulating sequence data and viewing structural representations of these molecules. A logical next step would be for students to explore phylogenetic relationships of these proteins. A key goal in this approach is to allow students to identify the strengths and weaknesses of disparate research approaches toward a more complete understanding of biological phenomenon. Equally important, students should begin to understand how these different approaches complement each other. Detailed majors module: Some plants produce proteins that can inhibit animal cells. One member of a group of such proteins, RIPs, (plant ribosome-inactivating proteins) is the cytotoxin ricin. This is one of the most powerful cytotoxins known and sometimes draws the attention of bioterrorists. Other such proteins act by destruction of the cell membrane. All of these plant toxins are probably part of a plant’s defense mechanisms. RIPs inactivate 60S ribosomal subunits by cleaving an N-glycosidic bond from a specific adenine base from the sugar-phosphate backbone of 28S rRNA. Toxins of this type include toxin from Shigella bacteria (shiga toxin), luffin (from gourds) and ricin (from castor bean). Many common grains also produce this kind of toxin. A distinctly different toxin, zeamatin, is produced by corn. This toxin destroys membrane permeability of cells and acts similar to alpha-amylase and trypsin inhibitor. Undergraduate ‘Wet” Laboratory A laboratory for isolating antifungal proteins from grains (Coleman, W., Bioscene 21:7-12, 1995; http://papa.indstate.edu/amcbt/volume_21/v21-2p7-12.pdf) affords student opportunities to experience classical protein purification procedures in introductory upper-level biochemistry laboratories. The laboratory incorporates aspects of microbiology, since protein inhibitory activity is assayed using a simple plate assay with a number of fungi. Students may select grains from different plants for which such protein activities have been observed. Further students may select other grains, tested or untested, for such activity. The assay was developed originally using Trichoderma reesei, but it may be used with Phycomyces blakesleeansis or commonly found yeast (Saccharomyces). Other fast growing fungi may be tested, also. The presence of these proteins during plant development and in parts of a mature plant might be investigated, also. This provides elements of inquiry not ordinarily found in such a standard, basic biochemistry laboratory. Instructors comfortable with this kind of laboratory may be introduced to the inquiry approach in a very non-threatening manner. Tritin contains 275 amino acids and has a molecular weight of 29613 daltons. This size is readily amenable to analysis using bioinformatics tools. This wet laboratory can be expanded to incorporate uses of data analysis tools for student inquiry. Students can discover if the gene for the protein isolated is present in other organisms. They may also find similarities/differences of their protein to those isolated by other class members. Are genes for the protein isolated present in other plants as well? What are the phylogenetic relationships of these proteins in these plants? Is the phylogenetic relationship of one protein (such as tritin) similar to that of ricin? Students will be introduced to these data analysis tools using the “Discovery Tree” from the CMS MBR site (http://restools.sdsc.edu/discovery_tools/discovery_tools.html). Procedures for students: 1. Introduction to the enzymes, their functions and sources 2. Give students names of the plants provided, for example, tritin is the name of the protein from wheat (Triticum aestivum). 3. You can find the sequence information for the protein you isolated by searching online databases such as SWISS-PROT. Use keywords such as tritin and Triticum, an accession number, or a specialized label will allow you to isolate the protein and its molecular information. The “Discovery Tree” provides a number of this and other such resources at the CMS MBR site. 4. Using these tools, find the DNA sequence of your protein. 5. Using BLAST, search for alignments of the sequence of your protein to that in other organisms. (Sequence homologies and other analyses) 6. Examine the protein for motifs that can help determine protein function. (Can use a variety of tools including PFAM. In particular, emphasize the domain nature of protein sequence/structure discussing domain swapping and assembly into new protein functions). 7. Using sequence information from similar proteins isolated in the class or provided by the instructor, perform a phylogenetic analysis.