| Fungal melanins | Despite their diverse functions as protective pigments and microbial virulence factors, the chemical structures of melanins have remained elusive because of their insolubility and amorphous nature. We are studying the molecular species involved in Cryptococcus neoformans melanin biosynthesis using selectively 13C-enriched obligatory exogenous precursors and NMR spectroscopy of both powdered and solvent-swelled melanin ‘ghosts’. In addition to the formation of cross-linked indole structures, we have found evidence for the presence of triglycerides in which we hypothesize the pigment is synthesized and for covalent linkages between the melanin and polysaccharide cell-wall components. |
| Cuticular biopolymers | Higher plants are encased in a cuticle that acts as a waterproofing layer and regulates the flow of nutrients among various plant organs. This project's goal is to understand the polymeric structures that make the outer skin of fruits resilient and resistant to pathogenic attack, especially under conditions of environmental stress. Using limes and tomatoes, studies are underway to find out how cutin interacts with waxes and water and to determine molecular structure for both intact cutin and its oligomeric fragments. Also under study are biosynthesis, structural development, and domain formation in suberin, a cuticular material that forms on the surface of wounded plant tissue. Finally, the work aims to determine the molecular identity of bbiopolymers responsible for hard-to-cook syndrome in potato tissue. This research program uses chemical and enzymatic methods to isolate and break down fruit cutins and suberins; structural studies are conducted using solution- and solid-state NMR, mass spectrometry, and atomic force microscopy. |
| Fatty acid-binding proteins | Fatty acids are important physiologically for energy storage and metabolism, membrane structure, and intracellular signal transduction. Fatty acid-binding proteins are small water-soluble proteins with well-characterized binding specificities and affinities for hydrophobic ligands but incompletely understood physiological function. Our group is studying the three-dimensional conformation and internal dynamics of fatty acid-binding proteins and associated complexes, including both wild-type proteins and site-directed mutants. This work employs a coordinated investigative strategy of multidimensional nuclear magnetic resonance (NMR), computational modeling, and fluorescence energy transfer assays. |
| NMR methodology application and development | Because many polymers and biopolymers are semisolid in nature, studies of their molecular structure and organization often require tailored spectroscopic approaches. First, NMR methodologies are under development to combine magic-angle spinning with multidimensional high-resolution NMR to yield quantitatively reliable and structurally informative spectra for lipid multilamellar dispersions and solvent-swelled plant cuticles. A second methodological initiative involves the application of two-dimensional wideline separation experiments to obtain motional profiles and spin diffusion rates in plant biopolymer and phospholipid membrane systems. Finally, solid-state REDOR and related experiments are being used to examine the three-dimensional molecular structure of peptide hormones, fungal melanins, and other amorphous biological solids. |
