Seminar Titles & Abstracts
Scientific Research Seminars
Hijacking pathogenic membrane proteins to engineer cellular entry: A molecular biophysics approach
Invasive pathogenic bacteria feature many cellular niches and life cycles, for which they have developed functions that are potentially attractive in biotechnology and therapeutic delivery applications. One such function is tissue-specific bacterial engulfment in human cells which do not normally undergo phagocytosis. Opacity-associated (Opa) proteins of Neisseria gonorrhoeae and N. meningitides are eight-stranded outer membrane proteins that bind to different host receptors, triggering engulfment of the bacterium. Receptor selectivity is determined by regions of the extracellular loops with high sequence variability; to date, 338 distinct opa alleles have been sequenced. Our hybrid NMR and molecular dynamics structure of an Opa reveals that the variable regions are dynamic and disordered, and sample conformations that may be competent for receptor binding. To complement our structural studies, Opa – human receptor interactions are being examined in vitro and in vivo, using liposomes and nanodiscs. In vitro studies focus on the strength and specificity of the interaction, while our in vivo studies are elucidating the cell fate and signaling of Opa liposomes. Our work suggests that a high degree of structural plasticity is required to tolerate the diverse sequences, and the dynamic ensemble allows sampling of conformers competent to engage receptors. Thus, perhaps counterintuitively, conformational plasticity offers a mechanism for functional versatility in this remarkable biological system.
Tuning Micelle Dimensions and Properties for Stabilizing Membrane Protein Fold and Function
One major bottleneck to the investigation of membrane proteins is the stabilization of structure and function in detergents and lipid bilayers after purification from the native membrane. Detergents are the most successful membrane mimic, thus far, for NMR and X-ray crystallography structure determination of membrane proteins. However, the current empirical screening of detergents that stabilize protein function and fold is laborious, costly, and often is not successful. To better understand the physical determinants that stabilize a protein-detergent complex, we have systematically investigated the properties of detergent micelles. Specifically, we have determined that binary detergent mixtures form ideally mixed micelles and that many physical properties are different from the pure individual micelles and vary linearly with micelle mole fraction. The predictability of the shape, size, and surface properties of binary mixtures expands the molecular toolkit for applications that utilize detergents and provide a means to systematically test the influence these properties have on membrane protein fold and function. Our progress towards correlating detergent micelle physical properties with membrane protein structure and function will be presented.
Membrane protein structural biology: from micelles to bacterial invasion
The research aims of the Columbus laboratory are two-fold: (1) to address the challenges in membrane protein structural biology and (2) to investigate the structural determinants of bacterial pathogen – host interactions mediated by membrane proteins. Specifically, the structures of N. gonorrhoeae and N. meningitides Opa proteins, which interact with host proteins and induce pathogen phagocytosis, are of interest. However, in order to investigate the structures of these proteins, as well as the complexes with the host receptors, methods for accelerating structure determination are being developed. A major obstacle to membrane protein structure determination is the selection of a detergent micelle that mimics the native lipid bilayer. Currently, detergents are selected by exhaustive screening because the effects of protein-detergent interactions on protein structure are so poorly understood. The overall goal is to use a multifaceted approach (NMR and EPR spectroscopy, small angle X-ray scattering, and X-ray crystallography) to develop a fundamental understanding of the protein – detergent interaction in order to accelerate the structure determination of membrane proteins involved in bacterial pathogenesis. Results arising from our current pursuits of this overarching, long-term goal will be presented.
Teaching and Research: a symbiotic relationship
Often the three pillars of academia (research, teaching, and service) are considered to be independent endeavors by faculty and administrators. This notion of mutual exclusivity likely stems from the finite time that an individual academic has to devote to each area. However, there is synergy between research, teaching, and service that can maximize time, improve student learning and training, and produce quality federally-funded research programs. In this presentation, I will present motivations and philosophies and developed programs that demonstrate the synergistic relationship of teaching and research. In addition to why I think this synergy is important to both student and faculty, I will specifically discuss an undergraduate biochemistry research-based laboratory curriculum, a research laboratory design that emphasizes learning and teaching for both graduate and undergraduate students, and how to incorporate experimental design and research into science curricula.