This project will have long-range impact on scientific understanding of a system of potential global importance. Many bacterial cells contain poorly understood interior structures called "microcompartments", which are composed of several thousand protein subunits, with a polyhedral outer shell reminiscent of viral capsids. The interior of these microcompartments is filled with enzymes carrying out sequential metabolic reactions important for life of the cell. They function as simple organelles in bacteria, which lack the well-studied membrane-bound organelles (e.g. mitochondria and chloroplasts) characteristic of cells from eukaryotes (e.g. animals and plants). The microcompartment under study in this project is the carboxysome. The function of the carboxysome is to enhance carbon sequestration and CO2 fixation. Microbes with carboxysomes make a major contribution to global CO2 fixation. An atomic level understanding of the carboxysome could be helpful in modifying a system that plays a major role in the global carbon cycle. Furthermore, related microcompartments in other bacteria are involved in producing various small organic molecules, some of which are relevant in biofuel applications. Initial structural studies on the protein shell of the carboxysome have led to hypotheses about how the protein shell might control the flow of metabolic intermediates into and out of the microcompartment. Research in the current project will engineer the carboxysome shell in order to advance understanding of these important and unique structures. The shell proteins will be engineered to modify the pores that are hypothesized to serve as gates for molecular transport into and out of the carboxysome. The modifications will test current hypotheses about molecular transport, which should lead to the ability to create microcompartments that can transport novel metabolites. In addition, the modifications will make it possible to isolate and study carboxysomes from model organisms amenable to genetic manipulation. Structural studies of large protein complexes like the carboxysome are at the leading edge of our abilities to investigate the molecular architecture of proteins.
Broader impacts: In addition to providing fundamental insight into large protein structures and mechanisms of production of reduced carbon in the biosphere, this project also involves a significant training component at both the graduate and undergraduate levels. The project will be executed by two graduate students with the assistance of a group of undergraduates drawn from programs geared to include underrepresented groups. This important training component of the project will have a substantial effect on education and scientific workforce development.
