In 1981, I came to MCW as a Muscular Dystrophy Association postdoctoral fellow, developing advanced electron paramagnetic resonance (EPR) spin labeling techniques to study membrane dynamics. I joined the MCW faculty in 1986.
We utilize site-directed spin labeling (SDSL) EPR spectroscopy and other biophysical and biochemical techniques to work on problems related to the membrane interactions of peptides and proteins and the structure and dynamics of membrane proteins.
Mechanism of Activation and Membrane Interactions of Pseudomonas toxin ExoU
ExoU is a 74 kDa protein produced by the Gram-negative bacterial pathogen, Pseudomonas aeruginosa. ExoU is injected directly into eukaryotic cells by a needle-like Type III secretion system (T3SS). Once inside the eukaryotic cell, ExoU functions as a phospholipase to disrupt membranes and facilitate dissemination of the bacterial infection. Activation of ExoU requires interaction with ubiquitin. Our site-directed spin label EPR studies in conjunction with NMR, mutagenesis, and biochemical analyses have provided an initial indication of the ubiquitin binding domain. Current aims of this research are to fully characterize the ExoU-ubiquitin binding interface and to understand how ExoU interacts with its target membrane substrate. These studies will facilitate the development of novel inhibitors of ExoU as a means to attenuate the virulence of P. aeruginosa infections.
Mechanisms of Antimicrobial Peptides: Membrane Interactions
Antibiotic resistance is an increasingly serious problem in the treatment of infectious disease. During the past two decades, a large number of peptides with potent antibacterial, antiviral, and antifungal properties have been identified from a wide range of both vertebrate and invertebrate species. These antimicrobial peptides (AMPs) are an important component of the innate arm of host resistance, serving as a first line of defense against infection. Despite being evolutionarily ancient, resistance to AMPs has only rarely been observed. Consequently, there is great interest in the development of these peptides for the treatment of drug-resistant infections.
Models of Transmembrane Channel Formation
(A) Peptide a-helices (cylinders) initially associate parallel to the membrane surface, either superficially (left) or embedded just below the aqueous interface. (B) Peptides continue to accumulate at or near the bilayer surface, disrupting lipid packing and causing membrane thinning. This step may or may not involve peptide-peptide aggregation. Once a critical peptide/lipid ratio is reached, peptides either (C) insert into the membrane as a barrel-stave type pore or (D) induce the localized formation of toroidal pores.
Our laboratory is involved in elucidating the mechanisms by which AMPs disrupt bacterial membrane structure, determining the basis of AMP selectivity for microbial cells, and developing more effective antimicrobial peptides and peptidomimetics. Whereas classical antibiotics generally target cell wall synthesis, protein translation, or some other highly specific target, AMPs are believed to function by directly disrupting the microbial cell membrane. Peptides are prepared using either recombinant DNA methods or solid-phase chemical synthesis, and their interactions with model membranes (liposomes) and cells are characterized by a variety of physical techniques including circular dichroism, fluorescence, and EPR site-directed spin labeling (SDSL). Our fundamental hypothesis is that a more complete understanding of peptide structure and dynamic interactions with the membrane will allow the design and development of improved AMPs and related antibiotics for the treatment of infections by existing drug-resistant strains such as Pseudomonas aeruginosa and methicillin-resistant Staphylococcus aureus (MRSA).
Samantha Kohn, Graduate Student
(Springer TI, Reid TE, Gies SL, Feix JB.) J Biol Chem. 2019 Dec 13;294(50):19012-19021 PMID: 31662432 PMCID: PMC6916483 SCOPUS ID: 2-s2.0-85076501851 10/31/2019
(Xia C, Shen AL, Duangkaew P, Kotewong R, Rongnoparut P, Feix J, Kim JP.) Biochemistry. 2019 May 14;58(19):2408-2418 PMID: 31009206 PMCID: PMC6873807 SCOPUS ID: 2-s2.0-85065650755 04/23/2019
(Feix JB, Kohn S, Tessmer MH, Anderson DM, Frank DW.) Cell Biochem Biophys. 2019 Mar;77(1):79-87 PMID: 30047043 PMCID: PMC6347562 SCOPUS ID: 2-s2.0-85050694302 07/27/2018
(Tessmer MH, Anderson DM, Pickrum AM, Riegert MO, Moretti R, Meiler J, Feix JB, Frank DW.) Proc Natl Acad Sci U S A. 2018 01 16;115(3):525-530 PMID: 29295930 PMCID: PMC5776994 SCOPUS ID: 2-s2.0-85042128224 01/04/2018
(Herneisen AL, Sahu ID, McCarrick RM, Feix JB, Lorigan GA, Howard KP.) Biochemistry. 2017 11 07;56(44):5955-5963 PMID: 29034683 PMCID: PMC6112238 SCOPUS ID: 2-s2.0-85033361759 10/17/2017
(Fischer AW, Anderson DM, Tessmer MH, Frank DW, Feix JB, Meiler J.) ACS Omega. 2017 Jun 30;2(6):2977-2984 PMID: 28691114 PMCID: PMC5494639 07/12/2017
(Tessmer MH, Anderson DM, Buchaklian A, Frank DW, Feix JB.) J Biol Chem. 2017 02 24;292(8):3411-3419 PMID: 28069812 PMCID: PMC5336173 SCOPUS ID: 2-s2.0-85013836015 01/11/2017
(Kim SS, Upshur MA, Saotome K, Sahu ID, McCarrick RM, Feix JB, Lorigan GA, Howard KP.) Biochemistry. 2015 Dec 15;54(49):7157-67 PMID: 26569023 PMCID: PMC4734095 SCOPUS ID: 2-s2.0-84949947657 11/17/2015
(Anderson DM, Feix JB, Frank DW.) PLoS Pathog. 2015 Jul;11(7):e1004944 PMID: 26203905 PMCID: PMC4512716 SCOPUS ID: 2-s2.0-84938811382 07/24/2015
(Anderson DM, Sato H, Dirck AT, Feix JB, Frank DW.) J Bacteriol. 2015 Feb;197(3):529-41 PMID: 25404699 PMCID: PMC4285982 SCOPUS ID: 2-s2.0-84920836378 11/19/2014
(Schultz KM, Feix JB, Klug CS.) Protein Sci. 2013 Nov;22(11):1639-45 PMID: 24123237 PMCID: PMC3831678 SCOPUS ID: 2-s2.0-84892738005 10/15/2013
(Del Alamo D, Tessmer MH, Stein RA, Feix JB, Mchaourab HS, Meiler J.) Biophys J. 2019 Dec 18 PMID: 31892409 01/02/2020