Antimicrobial peptides are evolutionarily highly preserved elements of the innate immune system in animals. They kill a wide range of microbial organisms such as bacteria, viruses, and fungi with high potency and speed. Their remarkable ability to prevent pathogenic resistance makes these peptides a viable alternative to conventional antibiotics. The broad objective of this research is to elucidate the structural basis for antimicrobial action, so that new and improved antibiotics with strong microbiocidal activity but low cytotoxicity to mammalian cells may be designed. ? ? The central hypothesis of this project is that antimicrobial peptides share common mechanisms of action that derive from the special characteristics of microbial membranes such as radius of curvature, surface charge, and lack of cholesterol, characteristics that are distinct from mammalian membranes. To test this hypothesis, we will specifically determine the interactions of two representative peptides with lipid bilayers that mimic bacterial, retroviral, and human erythrocyte membranes; determine the orientation topology of these peptides in these various lipid membranes; and measure the secondary structure and aggregation state of these peptides. The peptides of choice are protegrin-1 (PG-1) and rhesus theta-defensin 1 (RTD-1), which both possess a disulfide-bond stabilized beta-sheet conformation that is common to a large number of antimicrobial peptides, including the defensins found in humans. ? ? We will use an integrated solid-state nuclear magnetic resonance (NMR) approach to study the mechanism of action of these two beta-sheet peptides. The important lipid factors in antimicrobial selectivity will be identified by studying the lipid-peptide interactions in lipids with defined membrane curvature, cholesterol content, and anionic surface charges. 31P and 2H NMR will be used as the main probes for the lipid-peptide interaction. Information on the peptide orientation in the lipid bilayer is important for understanding whether the peptides disrupt the cell membrane by pore formation or by micellization. This information will be obtained by 13C and 15N NMR experiments using both oriented and unoriented static samples. The ability to extract molecular orientation using unoriented samples will allow us to measure the concentration-dependence and membrane-curvature-dependence of the peptide orientation. The secondary structure and aggregation of PG-1 and RTD-1 in lipid bilayers will be determined from NMR isotropic chemical shifts and multiple-quantum experiments, respectively. Together, the new structural information, correlated with the characteristics of lipid membranes, will significantly advance our understanding of the mechanism of action of beta-sheet antimicrobial peptides. Moreover, the proposed research will fill our knowledge gap of how beta-sheet peptides interact with lipid bilayers in general.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM066976-02
Application #
6695589
Study Section
Biophysical Chemistry Study Section (BBCB)
Program Officer
Chin, Jean
Project Start
2003-02-01
Project End
2008-01-31
Budget Start
2004-02-01
Budget End
2005-01-31
Support Year
2
Fiscal Year
2004
Total Cost
$223,786
Indirect Cost
Name
Iowa State University
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
005309844
City
Ames
State
IA
Country
United States
Zip Code
50011
Shcherbakov, Alexander A; Hong, Mei (2018) Rapid measurement of long-range distances in proteins by multidimensional 13C-19F REDOR NMR under fast magic-angle spinning. J Biomol NMR 71:31-43
Mandala, Venkata S; Williams, Jonathan K; Hong, Mei (2018) Structure and Dynamics of Membrane Proteins from Solid-State NMR. Annu Rev Biophys 47:201-222
Liao, Shu Y; Lee, Myungwoon; Hong, Mei (2018) Interplay between membrane curvature and protein conformational equilibrium investigated by solid-state NMR. J Struct Biol :
Kwon, Byungsu; Lee, Myungwoon; Waring, Alan J et al. (2018) Oligomeric Structure and Three-Dimensional Fold of the HIV gp41 Membrane-Proximal External Region and Transmembrane Domain in Phospholipid Bilayers. J Am Chem Soc 140:8246-8259
Roos, Matthias; Wang, Tuo; Shcherbakov, Alexander A et al. (2018) Fast Magic-Angle-Spinning 19F Spin Exchange NMR for Determining Nanometer 19F-19F Distances in Proteins and Pharmaceutical Compounds. J Phys Chem B 122:2900-2911
Gelenter, Martin D; Hong, Mei (2018) Efficient 15N-13C Polarization Transfer by Third-Spin-Assisted Pulsed Cross-Polarization Magic-Angle-Spinning NMR for Protein Structure Determination. J Phys Chem B 122:8367-8379
Lee, Myungwoon; Yao, Hongwei; Kwon, Byungsu et al. (2018) Conformation and Trimer Association of the Transmembrane Domain of the Parainfluenza Virus Fusion Protein in Lipid Bilayers from Solid-State NMR: Insights into the Sequence Determinants of Trimer Structure and Fusion Activity. J Mol Biol 430:695-709
Wang, Tuo; Jo, Hyunil; DeGrado, William F et al. (2017) Water Distribution, Dynamics, and Interactions with Alzheimer's ?-Amyloid Fibrils Investigated by Solid-State NMR. J Am Chem Soc 139:6242-6252
Lee, Myungwoon; Wang, Tuo; Makhlynets, Olga V et al. (2017) Zinc-binding structure of a catalytic amyloid from solid-state NMR. Proc Natl Acad Sci U S A 114:6191-6196
Fritzsching, Keith J; Hong, Mei; Schmidt-Rohr, Klaus (2016) Conformationally selective multidimensional chemical shift ranges in proteins from a PACSY database purged using intrinsic quality criteria. J Biomol NMR 64:115-30

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