Collaborative Research: A Comprehensive Study of the Structure, Function, and Diversity of Detoxification Enzymes (CYP2B) in Mammalian Herbivores (Neotoma)

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At every meal, thousands of species of plant-eating mammals confront the possibility of being poisoned by the natural toxins produced by their food. The generalized foraging habit of mammalian herbivores, eating many different plant species on a daily basis, is thought to result from limitations of their liver enzymes. Generalists are hypothesized to have a "jack of all trades" style of detoxification system that consists of liver enzymes able to metabolize a wide array of plant toxins but with low catalytic efficiency. In contrast, specialists are thought to have detoxification enzymes with enhanced efficiency towards a restricted number of toxins. This concept of biochemical tradeoffs with respect to dietary specialization has essentially become dogma without any experimental support. This research will capitalize on the extensive knowledge of mammalian detoxification enzymes gained from studies of drug metabolism in model species and humans, as well as on the availability of a superlative wild rodent model of mammalian herbivory. The long-term objective is to determine how dietary exposure to plant toxins has shaped the mammalian detoxification system through a focused study of liver enzymes in the cytochrome P450 subfamily 2B (CYP2B) in herbivorous woodrats (Neotoma). The specific goals of this collaborative research are: 1.) Characterize the CYP2B enzymes in woodrats at the amino acid level. 2.) Use new metabolomic approaches to identify key P450 substrates in diets of woodrats and compare metabolism of plant toxins by woodrat CYP2B enzymes. 3.) Conduct structure-function analyses using site-directed mutagenesis and X-ray crystallography of woodrat CYP2B enzymes. The work will address a longstanding hypothesis by drawing upon cutting-edge biochemical and structural biology approaches to generate the first detailed examination of CYP2B enzymes of wild herbivores, identify substrates, and compare enzyme function. The ultimate goal is to solve the crystal structures of woodrat enzymes in complex with plant toxins. Currently, knowledge of detoxification enzymes of mammals is rudimentary. The work has the potential to contribute to the understanding of drug metabolism in humans given preliminary data of these investigators on woodrat enzymes. In addition, woodrats are nature's historians; their behavior of storing food has facilitated an understanding of the natural changes that have taken place in the desert southwest over the last 35,000 years. An interactive display about the ecology of woodrats and their importance to society will be developed at the Utah Museum of Natural History. The PIs will mentor a new generation of trainees (high school students through postdoctoral fellows) in interdisciplinary research. This award is co-funded by the Chemistry of Life Processes Program in the Division of Chemistry.