Mechanism of action of multi-hit host defense peptides: Molecular basis for anti-infective and immunomodulatory properties.
- This ongoing project is currently funded by the National Science Foundation (MCB 1716608) and National Institutes of Health:
- We investigate piscidins as host defense (antimicrobial) metallopeptides active on a broad range of bacteria, including C. difficile. Using methods that include CD- and NMR-monitored titrations, structural studies by SSNMR and neutron diffraction, surface plasmon (SPR), quartz crystal microbalance (QCM), calorimetry, microscopy, gel retardation assays, Ca2+-release assays, and in vitro assays for determination of cytotoxicity and cell death mechanisms, we focus on identifying the cellular targets of several isoforms and characterizing how targets are disrupted by peptides in the context of living cells and their mimics.
- Our new investigations show that piscidins bound to Cu act as nucleases that damage bacterial DNA and as lipoxygenases that oxidize unsaturated fatty acids. They work synergistically with Cu to kill bacteria (including biofilms) and cancer cells. Specifically, the peptides use an amino terminal Cu and Ni (ATCUN) motif (XXH; X = variable residue) to mediate Cu(III)/Cu(II) redox cycles and generate ROS in the presence of Cu and co-reactants. Metal binding also improves their membrane capability.
Featured publications include:
- R. Fu, M.T. Rooney, R. Zhang, and M.L. Cotten. Coordination of Redox Ions within a Membrane-Binding Peptide: A Tale of Aromatic Rings. 2021. J. Phys. Chem. Letters. https://doi.org/10.1021/acs.jpclett.1c00636.
- F. Comert, F. Heinrich, A. Chowdhury, M. Schoeneck, C. Darling, K.W. Anderson, M.D.J. Libardo, A.M. Angeles-Boza, V. Silin, M.L. Cotten, and M. Mihailescu. Copper-binding anticancer peptides from the piscidin family: an expanded mechanism that encompasses physical and chemical bilayer disruption. Just accepted, Sci. Reports. https://www.nature.com/articles/s41598-021-91670-w.
- Paredes, S. Kim, M.T. Rooney, A.I. Greenwood, K. Hristova, and M.L. Cotten. Effect of Metallation on the Membrane Activity of Piscidin: Mechanistic Insights Gained from 31P Solid-state NMR. Invited manuscript to Biochim. Biophys. Acta – Biomembranes, Commemorative Special Issue in Honor of Michèle Auger. 2020, 1862:183236. https://doi.org/10.1016/j.bbamem.2020.183236.
- Cetuk, J. Maramba, M. Britt, R. Ernst, E. Mihailescu, M.L. Cotten and S. Sukharev. Differential interactions of Piscidins with phospholipids and lipopolysaccharides at membrane interfaces. Langmuir, April 19, 2020, https://doi.org/10.1021/acs.langmuir.0c00017.
- Juliano, L. Serafim, S. Duay, M. Heredia Chavez, G. Sharma, M. Rooney, F. Comert, A. Radulescu, M.L. Cotten, M. Mihailescu, E. May, A. Greenwood, R. Prabhakar, A. Angeles-Boza. A Potent Host Defense Peptide Triggers DNA Damage and is Active against Multi-Drug Resistant Gram-Negative Pathogens. ACS Infectious Diseases, April 6, 2020, https://doi.org/10.1021/acsinfecdis.0c00051.
M. Mihailescu, M. Sorci, J. Seckute, V. I. Silin, J. Hammer, B. Scott Perrin Jr., J. I. Hernandez, N. Smajic, A. Shrestha, K. A. Bogardus, A. I. Greenwood, R. Fu, J. Blazyk, R. W. Pastor, L. K. Nicholson, G. Belfort, M. L. Cotten. Structure and Function in Antimicrobial Piscidins: Histidine Position, Directionality of Membrane Insertion, and pH-Dependent Permeablization. 2019. J Am Chem Soc. 141:9837-9853.
- Comert, A.I. Greenwood, J. Maramba, R. Acevedo, L. Lucas, T. Kulasinghe, L.S. Cairns, Y. Wen, R. Fu, J. Hammer, J. Blazyk, S. Sukharev, M.L. Cotten, and M. Mihailescu. The Host-defense Peptide Piscidin P1 Reorganizes Lipid Domains in Membranes and Decreases Activation Energies in Mechanosensitive Ion Channels. 2019. J. Biol. Chem. 294:18557-18570.
- Oludiran, D.S. Courson, M.D. Stewart, M.L. Cotten, and E. B. Purcell. How Oxygen Availability Affects the Antimicrobial Efficacy of Host Defense Peptides: Lessons Learned from Studying the Copper-binding Peptides Piscidins 1 and 3. 2019. Int J Mol Sci. 20(21).
R.K. Rai, A. De Angelis, A. Greenwood, S.J. Opella, M. L. Cotten. Metal-ion Binding to Host Defense Peptide Piscidin 3 Observed in Phospholipid Bilayers by Magic Angle Spinning Solid-State NMR. 2019. Chemphyschem. 20:295-301.
S.Y. Kim, F. Zhang, W. Gong, K. Chen, K. Xia, F. Liu, R. Gross, J.M. Wang, R.J. Linhardt, and M.L. Cotten. Copper Regulates the Interactions of Antimicrobial Piscidin Peptides from Fish Mast Cells with Formyl-peptide Receptors and Heparin. 2018. J. Biol. Chem. 293:15381-15396.
- M.D.J Libardo, A.A. Bahar, B. Ma, R. Fu, L.E. McCormick, J. Zhao, S.A. McCallum, R. Nussinov, D. Ren, A.M. Angeles-Boza, and M.L. Cotten. Nuclease activity gives an edge to host-defense peptide piscidin 3 over piscidin 1, rendering it more effective against persisters and biofilms. 2017. FEBS J. 284:3662-3683
- R.M. Hayden, G.K. Goldberg, B.M. Ferguson, M.W. Schoeneck, M.D.J. Libardo, S.E. Mayeux, A. Shrestha, K.A. Bogardus, J. Hammer, S. Pryshchep, H.K. Lehman, M.L. McCormick, J. Blazyk, A.M. Angeles-Boza, R. Fu, and M.L. Cotten. Complementary Effects of Host Defense Peptides Piscidin 1 and Piscidin 3 on DNA and Lipid Membranes: Biophysical Insights into Contrasting Biological Activities. 2015. J. Phys. Chem. B. 119:15235-46. We provided the first characterization of DNA/peptide aggregates by solid-state NMR and showed complementary effects between piscidin isoforms. We collaborated with groups at Hamilton College and the University of Connecticut.
- B.S. Perrin Jr., Y. Tian, R. Fu, C.V. Grant, E.Y. Chekmenev, W.E. Wieczorek, A.E. Dao, R.M. Hayden, C.M. Burzynski, R.M. Venable, M. Sharma, S.J. Opella, R.W. Pastor, and M.L. Cotten. High-Resolution Structures and Orientations of Antimicrobial Peptides Piscidin 1 and Piscidin 3 in Fluid Bilayers Reveal Tilting, Kinking, and Bilayer Immersion. 2014. J. Am. Chem. Soc. 136: 3491–3504. Cover story, podcast, and spotlight entitled “Solid-State NMR Reveals How a Kinky Peptide Slays Microbes”. We solved the first high resolution structures of alpha-helical host defense peptides bound to fluid bilayers and deposited them in the Protein Data Bank. This work featured a collaboration with Dr. Rich Pastor at the NIH.
- R. Fu, E.D. Gordon, D.J. Hibbard, and M. Cotten. High resolution heteronuclear correlation NMR spectroscopy of an aligned antimicrobial peptide: Direct evidence of spectral enhancement at high magnetic field and peptide-water interactions at the water-bilayer interface. 2009. J. Am. Chem. Soc. 131: 10830–10831. We collected NMR data on state-of-the-art instruments, including the 900 MHz ultra-wide bore spectrometer at the National High Magnetic Field Laboratory, and helped develop new solid-state NMR methodology for membrane-bound species. These experiments were done in collaboration with Dr. Riqiang Fu at the NHMFL.