Poulter Research Group

Research

My research group is interested in problems at the interface between organic chemistry and biochemistry. A major focus is the reactions catalyzed by enzymes in the isoprene biosynthetic pathway with special emphasis on establishing the mechanisms of the enzyme-catalyzed transformations and how the enzymes promote the reactions. Another area of interest is structure-function relationships in nucleic acids, in particular the topologies of complex naturally occurring RNAs, how their shapes relate to biological function and what governs their interactions with other biopolymers during protein biosynthesis.

The isoprene biosynthetic pathway is needed by all organisms to produce essential compounds. We are studying several key enzymes in the pathway that catalyze fundamental reactions, including the isomerization of isopentenyl diphosphate (IPP) to dimethylallyl diphosphate (DMAPP), the condensation of IPP with a variety of allylic diphosphates to yield new allylic isoprenylogues containing five additional carbons, and the unusual rearrangement of presqualence diphosphate (PSPP) to squalene during bio-synthesis of cholesterol. We are also studying unusual isoprenoid alkylations that occur during biosynthesis of ergot alkaloids, post-translational modifications of proteins, and in transfer RNAs. We isolate genes for many of the enzymes we study, construct plasmids for overexpression of the enzymes, and conduct site-directed mutagenesis on critical amino acids to elucidate their role in catalysis.

We work in collaboration with X-ray crystalographers to obtain 3-dimensional structures for isoprenoid enzymes, and use these structures to alter the proteins to better understand how they function as catalysts. All of the reactions presented above are interesting biological alkylations in that they do not rely on commonly observed carbonyl group chemistry for construction of carbon carbon and carbon-heteroatom bonds. Work in this area provides training in a combination of biochemical (purification of enzymes, kinetics, precursor-product studies) and chemical (synthesis, isolation-identification, reaction mechanisms) techniques.

Fish-hunting cone snails from tropical waters immobilize their prey by injecting a venom that contains a rich mix of small peptide toxins. These molecules are among the most selective and potent compounds known to target a whole host of cellular receptors. We are using high field NMR to determine the structures of the cone snail peptide toxins at high resolution in order to deduce what factors are important for their highly selective binding. These proteins are relatively small (~10 to 30 amino acids), and are well suited to analysis by multidimensional NMR techniques.

Much of what we do relies heavily on modern analytical methods such as high pressure liquid chromatography, high field multinuclear magnetic resonance spectroscopy, and gas chromatography-mass spectrometry. We draw on ideas and techniques from organic chemistry and biochemistry to design our experiments, including developing new procedures for synthesis of compounds, developing new methods for measuring interactions between an enzyme and its substrates, and using recombinant DNA technology to obtain biological molecules that are normally difficult to isolate. It is the interdisciplinary nature of our work that we find both challenging and exciting.