NMR spectroscopy will be developed as a method for structural studies of proteins in supramolecular structures and applied to the filamentous bacteriophages, especially phages with coat proteins displaying designed and selected peptides on their surface. The research plan for the next funding period reflects the success in developing solid-state NMR spectroscopy for the study of proteins achieved with the support of this grant, the greatly improved capabilities of a newly constructed high-field solid-state NMR spectrometer dedicated to these studies, and recent developments in the molecular biology of filamentous bacteriophages, which enable them to display peptide sequences inserted into the major pVIII coat protein. The major area of spectroscopic development continues to be solid-state NMR method for determining the structures of proteins. The approach receiving the greatest attention is for samples where the proteins are immobile and uniaxially oriented in the magnetic field of the spectrometer. Complementary spectroscopic methods will be developed which utilize magic angle sample spinning on samples where the proteins are immobile and unoriented. Methods for using solid-state NMR spectroscopy to characterize the backbone dynamics of proteins will also be developed. Motional averaging of powder pattern lineshapes will be supplemented with relaxation measurements. Molecular dynamics simulations with and without experimental restraints are an integral part of the research plan. The proposed studies exploit several unique biological and physical properties of filamentous bacteriophages, especially the folding and immobilization of the inserted peptide sequences in the environment of the coat proteins and the spontaneous orientation of the phages in the magnetic field of the NMR spectrometer. Studies of these peptides are of interest from the perspectives of protein folding, epitope characterization, and receptor binding. Peptide sequences important to understanding diseases as diverse as malaria, cancer, and H.I.V. will be investigated.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
4R37GM024266-22
Application #
2468076
Study Section
Special Emphasis Panel (NSS)
Project Start
1977-08-01
Project End
2003-07-31
Budget Start
1998-08-01
Budget End
1999-07-31
Support Year
22
Fiscal Year
1998
Total Cost
Indirect Cost
Name
University of Pennsylvania
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Sinha, Neeraj; Grant, Christopher V; Wu, Chin H et al. (2005) SPINAL modulated decoupling in high field double- and triple-resonance solid-state NMR experiments on stationary samples. J Magn Reson 177:197-202
Nevzorov, Alexander A; Mesleh, Michael F; Opella, Stanley J (2004) Structure determination of aligned samples of membrane proteins by NMR spectroscopy. Magn Reson Chem 42:162-71
Mesleh, M F; Valentine, K G; Opella, S J et al. (2003) Myristoylation as a general method for immobilization and alignment of soluble proteins for solid-state NMR structural studies. J Biomol NMR 25:55-61
Nevzorov, Alexander A; Opella, Stanley J (2003) A ""magic sandwich"" pulse sequence with reduced offset dependence for high-resolution separated local field spectroscopy. J Magn Reson 164:182-6
Mesleh, Michael F; Lee, Sangwon; Veglia, Gianluigi et al. (2003) Dipolar waves map the structure and topology of helices in membrane proteins. J Am Chem Soc 125:8928-35
Park, Sang Ho; Mrse, Anthony A; Nevzorov, Alexander A et al. (2003) Three-dimensional structure of the channel-forming trans-membrane domain of virus protein ""u"" (Vpu) from HIV-1. J Mol Biol 333:409-24
Zeri, Ana Carolina; Mesleh, Michael F; Nevzorov, Alexander A et al. (2003) Structure of the coat protein in fd filamentous bacteriophage particles determined by solid-state NMR spectroscopy. Proc Natl Acad Sci U S A 100:6458-63
Lee, Sangwon; Mesleh, Michael F; Opella, Stanley J (2003) Structure and dynamics of a membrane protein in micelles from three solution NMR experiments. J Biomol NMR 26:327-34
Mesleh, Michael F; Opella, Stanley J (2003) Dipolar Waves as NMR maps of helices in proteins. J Magn Reson 163:288-99
Marassi, Francesca M; Opella, Stanley J (2003) Simultaneous assignment and structure determination of a membrane protein from NMR orientational restraints. Protein Sci 12:403-11

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