Current projects in the Cremeens Lab focus on one basic question in the context of two very different projects - “How does ‘it’ work, what is the mechanism?”. Please see below for more detail.
Biophysical Chemistry Project:
“Mechanistic studies of bacterial cell membrane-peptide interactions”
Knowing how antimicrobial peptides interact with bacterial membranes is important for their potential development as therapeutics. Most studies attempting to address antimicrobial mechanisms of peptides employ model membranes; however, the complexity of cell membranes causes concern about the degree to which conclusions created using model membranes can be extended to bacterial cell membranes. A membrane-disruption mode of antimicrobial activity would be sensitive to severe truncations of the peptide and would inhibit growth as well as be bactericidal. If inhibitory and bactericidal concentrations appear to be sensitive to the specific bacterial strains employed, as they appear to be in our preliminary results, then directly comparing membrane lipid composition for each strain will be important for assessing membrane-disruption versus non membrane disruption modes of action. The research team’s goal is to evaluate the potential for a membrane-disruption mode of action for antimicrobial peptides, as opposed to non-membrane-disruption mode of action, through biophysical characterization of peptide-lipid interactions, biological characterization of peptide-bacteria interactions, and the characterization of bacterial membrane lipid and peptidoglycan composition.
Physical Organic Chemistry Project:
“Assessing the Possibility and Probability of Surface Crossings in High Energy Ring Expansions”
Commonly, thermal organic reaction mechanisms have focused on ground-state electronic surfaces. In what non-photochemical scenarios ought an organic chemist take seriously the possibility of thermally accessing excited-state electronic surfaces? Upon going from closed-shell reactants to open-shell intermediates, non-adiabatic paths to excited-states are possible. The reaction pathways of nine fused cyclopropane derivatives were mapped using density functional theory (DFT) and complete active space self-consistent field (CASSCF) methods. Since these ring openings involve relatively high-energy species that lead to low energy aromatic species, a common scenario for non-adiabatic reaction paths, we posited that such reaction paths might come close to, or cross, excited-state surfaces. Although all nine cases showed the possibility of surface crossings, only one had a reported synthesis, 6-methylidenebicyclo[3.1.0] hex-3-en-2-one. Efforts are underway to synthesize this molecule using a three-step synthesis outlined by Rule et al. (Tetrahedron 1982 787). Experimentally, we aim to differentiate between excited-state zwitterions and ground-state triplet biradical intermediates in trapping studies, whereby products from heterolytic and homolytic reactions will be correlated with zwitterionic and biradical states, respectively. The overall goals of the project are to (1) computationally characterize a potentially non-adiabatic ring-opening reaction, (2) synthesize ring-strained molecules to experimentally test the predictions, and (3) experimentally investigate the ring-opening mechanisms.
Relevance: Incomplete combustion of petroleum leads to soot formation by a mechanism that involves polycyclic aromatic hydrocarbons. Since combustion involves relatively high-energy intermediates that lead to low-energy molecules (i.e. aromatic hydrocarbons), a common scenario for non-adiabatic reaction paths, it is plausible that combustion reaction paths might involve excited state surfaces or non-adiabatic events. The potential generality and relevance of non-adiabatic reaction paths in combustion is yet to be thoroughly explored. New mechanistic insights for soot formation could facilitate a paradigm shift in reducing soot formation.
Methods: Synthesis, Spectroscopy, and Calculations
Small organic molecules for the physical organic chemistry project are synthesized via traditional organic synthesis, i.e. using inert atmosphere techniques, thin layer chromatography (TLC), column chromatography, gas chromatography—mass spectrometry (GC-MS), and nuclear magnetic resonance (NMR) spectroscopy. Peptides are synthesized by solid-phase peptide synthesis, purified by HPLC, and characterized by electrospray ionization mass spectrometry (ESI-MS), circular dichroism (CD), infrared spectroscopy (IR), and/or NMR spectroscopy. For some biophysical characterizations, site-specific carbon-deuterium (C-D) or nitrile (C≡N) labeled peptides will be synthesized and investigated via IR spectroscopy.
Experimental observations are often compared to quantum chemical calculations using Gaussian and desktop workstations or via GAMESS, NWCHEM and the WebMO interface on a cluster at Gonzaga University’s Intel Corporation Computational Science Laboratory.
A collaboration with the Corcelli Lab at Notre Dame provides additional theoretical insights into our spectroscopic observations. Raman spectroscopy is also used to explore our systems via a collaboration with the Desamero Lab at York College.
Support comes from Gonzaga University start-up funds, the Gonzaga Science Research Program, Gonzaga’s Department of Chemistry and Biochemistry, and in part by a grant to Gonzaga University from the Howard Hughes Medical Institute through the Undergraduate Science Education Program.
Acquisition of a CD spectrometer was supported by the National Science Foundation under CHE-0922945 (“MRI: Acquisition of a Spectropolarimeter: A Chiro-Optical Spectroscopy Workbench” $181,155; 2009-2012; PI: Cremeens with co-PIs Jeff Watson and Tommaso Vannelli.)