Maria D. Vitery, Biochemistry and Molecular Biology
Crotamine, the Protein from the Venom of the South American Rattlesnake, and its Binding to DNA
Faculty Mentor: Dr. Richard Karpel
Crotamine is a protein from the venom of the South American rattlesnake (Crotalus durissus terrificus). This 42-residue polypeptide is a nucleic acid binding protein that is capable of penetrating cells and targeting chromosomes. It has the ability to carry plasmid DNA into cells that are actively proliferating. Cell penetration is believed to follow interaction of crotamine with cell surface heparan sulfate proteoglycans. This quality makes crotamine a potential candidate for drug transport. The goal in the lab is to quantify salt dependence, binding site size and affinities of crotamine for DNA. Our experiments focus on studying the binding of crotamine to single- and double-stranded DNA over different ionic conditions. In these experiments, the fluorimeter is used to determine light scattering, which is a measure of particle size. In parallel, we perform experiments that include the usage of a DNA-intercalating dye, ethidium bromide. Ethidium bromide intercalates within DNA and absorbs light at 600nm, and becomes fluorescent when bound to DNA. In these experiments, the interruption of DNA and ethidium bromide fluorescence by crotamine and its reversal by heparin is being quantified.
When and how did you find out that you could do independent research or creative work as a UMBC undergraduate?
I found out about the independent research at UMBC during my first semester after I transferred from Montgomery College.
How did you find a mentor and decide on a project?
I have worked on protein (molecular chaperones) at the NIH, therefore when I transferred to UMBC I wanted to keep doing research on proteins. I went on the Department of Chemistry and Biochemistry website and read about the different Principal Investigators and the research they perform. I really liked the research of Dr. Karpel so I decided to request an interview with him.
How did you know this was the project you wanted to do?
Crotamine was discovered fairly recently, and there is not much information about its properties and chemical characteristic yet. Having the chance of working on a "new" protein and finding the its binding affinities to DNA and RNA in different conditions, made me realize that this was the project for me. All the findings that I would obtain would help the characterization of crotamine and in future use as a drug-delivery vehicle against actively proliferating cells.
How much time did you put into it?
I try to take the research I do at the lab very seriously. Normally, I go four to five times a week for 4 to 5 hours. Characterizing a new protein is very interesting and fascinating since you do not have old data or previous information about it. The downside to this is that setting up the experiments is very time consuming and sometimes you do not obtain any results after you repeat the experiment several times. Therefore, it is required to spend a lot of time working on each experiment.
What academic background did you have before you started?
As I mentioned before, I have work at the NIH National Cancer Institute for two summers. There I also worked on proteins, so I had background on protein purification and characterization. Also, I have taken several upper level biochemistry and molecular biology courses that made me capable of performing this independent research.
What has been the hardest part about your research/creative work? The most unexpected thing?
The most unexpected thing was spending more the three months on only repeating one experiment. Recently I have been working on quantifying the decrease in fluorescence of crotamine upon addition of plasmid DNA. At first when I set up this experiment it seemed pretty straight forward, but this was not easy. It has taken me more than three months to perfect the protocol for this experiment and to be able to get reproducible data.
How does your research/creative work relate to your work in other classes?
As a Biochemistry and Molecular Biology major, I learn about different major proteins in most of my upper level courses. This project helps me to put all the knowledge and theory learned in my classes into practice.
What is your advice to other students about getting involved in research?
My best advice is do not be afraid to get involved in a independent research project. Even if you do not get the outcomes that you expected, I am pretty sure that at the end you will still have learned something very valuable.
What are your career goals?
I am planning on going into an M.D/Ph.D after I graduate, and do research in neuroscience.
How did you decide to present your work at URCAD this year?
URCAD is a great way to get to know people that work in the same area that you do and learn from their exeripences. Also, I think it is a good way to show my fellow classmates what I have achieved after a year of independent research.
Was the application difficult to complete? How much did your mentor help you with the application?
The application was not difficult, but it was time consuming. Writing a good abstract requires a lot of time and dedication. Dr. Karpel, was always willing to write my rough draft and give me as much feedback as possible.