The objective of this collaborative project is to use fluorescence energy transfer to recover the distance distributions between sites on peptides of known sequence. Studies of distance distribution in proteins, membranes and other biochemical systems is actively being investigated under GM- 35154, which is funded until July, 1994. In the present application, we propose to study the conformational distributions of helix-forming peptides using fluorescence resource energy transfer (FRET) and frequency-domain fluorometry. The peptides will be labeled with donors (D) and acceptors (A), typically and DANSYL, respectively. Additionally, we have available a range of acceptors for tryptophan with Forster distances ranging from 8 to 35Ao. The specific peptides will include the S-peptide from ribonuclease, which will be examined in helix-forming solvents and under denaturing conditions. The donor will be placed centrally, with an acceptor on either end, to test whether one-half of the peptide preferentially forms a helix. Following initial studies, electrostatic effects will be introduced by changes in the sequence to reveal the effects of these interactions on helix formation. Frequency-domain studies of trp-to-acceptor energy transfer will be used to determine the extents of helix formation in each half of the peptide. We will attempt to determine whether helix formation occurs in a one-step or continuous manner. Additionally, trp-to-acceptor diffusion coefficients will be obtained from the analysis, and should reveal the dynamics of these peptides in aqueous solutions. Additional bioactive peptides with donors and acceptors will include the neuropeptide galanin (GAL), gonadotropin releasing hormone (GnRH), somatostatin (SST) and calcitonin (CT). The role of Professor Kupryszewski's laboratory is to synthesize and purify the peptides, which will be designed jointly by Drs. Kupryszewski and Lakowicz. The synthetic effect is not supported by the FIRCA parent grant GM-35154.

Agency
National Institute of Health (NIH)
Institute
Fogarty International Center (FIC)
Type
Small Research Grants (R03)
Project #
5R03TW000049-02
Application #
3432463
Study Section
Special Emphasis Panel (SRC)
Project Start
1992-04-01
Project End
1995-03-31
Budget Start
1993-04-01
Budget End
1994-03-31
Support Year
2
Fiscal Year
1993
Total Cost
Indirect Cost
Name
University of Maryland Baltimore
Department
Type
Schools of Medicine
DUNS #
003255213
City
Baltimore
State
MD
Country
United States
Zip Code
21201