The purpose of this project is to use primarily x-ray crystallography to study structure-function relationships in heme containing enzymes and proteins. So far the focus has been on peroxidases. Peroxidases participate in a wide range of biological processes requiring oxidatiion of molecules and removal of peroxides. Several peroxidase structures have been solved so the project will now move into a more detailed analysis of function. In particular, the focus will be on chloroperoxidase (CPO), the most diverse of the known heme enzyme catalysts. The goal is to couple mutagenesis, molecular modeling, and crystallography in order to understand how CPO catallyzes stereoselective epoxidation, sulfoxidtion, and hydrogen abstraction reactions. In addition, the structure of the novel heme catalyst, heme oxygenase, will be determined. Recently, it has been found that some peroxidase crystals when frozen in liquid nitrogen diffract to beyond 1A resolution using synchrotron radiation. This provides a rare opprtunity for determining the structure of moderately large glycoproteins at true atomic resolution. Studies also will be initiated on novel heme proteins that operate as biological sensors and transcription factors. A transcriptional regulatory protein that senses carbon monoxide will be a particular focus.
The enzymes under investigation catalyze a variety of biologically important oxidations. A common theme is the presence of a heme prosthetic group. The remarkable feature in these proteins is the wide range of functions even though the hene group is exactly the same. These functions include oxygen carriers (hemoglobin and myoglobin), electron transfer proteins (cytochromes), and the enzymes under investigation in this project which catalyze bond making and breaking reactions. Some heme enzymes catalyze reactions that are important for degrading toxic environmental pollutants as well as stereospecific oxidations. Such reactions are quite important in various industrial processes where the need for stereoselectivity is high. Engineeering enzymes for this purpose is an area of considerable interest in thr synthesis of rare and expensive intermediates used in the production of important organic molecules. In recent years, a number of these important enzymes have been cloned and expressed in bacterial hosts. This opens the way for engineering these enzymes to tailor them for specific functions. However, before such engineering efforts can move forward, it is necessary to determine the three dimensional structural. Achieving this goal of structue determination using x-ray crystallography is the primary focus of this research.
