Investigating Molecular Recognition and Biological Function at Interfaces Using Antimicrobial Peptides.
EMSL Project ID
10595b
Abstract
This research proposal features fundamental principles underlying biological function and mechanisms of action of peptides active at interfaces. The strategy for this project has been to use complementary solid-state Nuclear Magnetic Resonance (ssNMR) techniques on site-specific isotopically labeled peptides. A number of circular dichroism (CD) and solution NMR experiments have also been planned to help guide the ssNMR work. Syntheses, purification, and identification of the key peptides as well as their preparation for NMR experiments can and have been performed at Pacific Lutheran University (PLU, Tacoma, WA, http://www.plu.edu/). In order to permit progress towards the ssNMR-related goals of this proposal, we are applying for blocks of time on the EMSL 500 WB Varian Unity+ and 600 NB Varian Unity NMR spectrometers. Note that our work, which started in the summer of 2003 with the use of Magic Angle Spinning techniques (REDOR, see below) to obtain distance restraints, is now ready to phase in a complementary technique based on the use of oriented samples to get orientational restraints. It is therefore an exciting time for this project. I would also like to mention that our results collected at the EMSL were featured in a talk, which I was invited to give at an American Chemical Society Symposium on solid-state NMR (Great Lakes Regional Meeting, Peoria, October 2004). Another invited talk has been scheduled for September 2005, at a Symposium on "Antimicrobial Peptides and Membrane Biophysics" in Ann Harbor, Mi. So, if the core of our proposal has retained many of its original features, we have clearly made significant progress in achieving our initial goals and the future work proposed here is promising.As part of our plans, the mechanisms of action of piscidins, antimicrobial peptides from mast cells of fish, have been studied through the investigation of structure/dynamics/function relationships. These topics are addressed by a combination of robust ssNMR techniques. More precisely, we have successfully performed REDOR distance measurements (13C to 15N) on labeled peptides interacting with hydrated lipid bilayers. In order to obtain orientational constraints from oriented samples we have purchased two flat coil probes (31P/1H and 15N/1H) through funds recently received from Research Corporation. Such probes are not available at the EMSL at this point in time. We were very enthusiastic to test the 31P probe at the EMSL for a few days in March 2005. We are presently taking advantage of our time allotment for the period of April-October 2005 and we have successfully started the investigation of oriented peptide-lipid samples. Overall, we have made considerable progress towards our goals of characterizing the peptide structure and orientation in lipid bilayers. Hence, we are working on the draft of a manuscript (featuring Figure 4 and Table 3) which we plan to submit to a peer-reviewed journal this summer. We are applying for additional NMR time during the period of October 2005-April 2006 so that we can complete the final goals outlined in our Research Corporation proposal. This grant will be considered for renewal within the next year so it is important for us to meet these remaining goals. The incoming work will involve the dynamics studies using 15N-backbone labeled peptides. The experiments will be performed at several temperatures.
Understanding molecular recognition at interfaces, as proposed here, can provide some important knowledge needed to fight many diseases and design new drugs, including broad-spectrum ones. Beyond this scientific motivation, a paramount goal of this project is also to provide an environment in which undergraduate students can integrate learning and advanced research.
Access to the state of the art facility at EMSL would be pivotal to achieving my pursuits, both from research and educational standpoints. Thank you for your consideration.
Project Details
Project type
Capability Research
Start Date
2005-12-19
End Date
2006-11-21
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members