Heme is an important biological compound composed of an iron atom inserted into a porphyrin by the enzyme ferrochelatase. Heme can be covalently linked to a protein by another enzyme called holocytochrome c synthase (HCCS) to generate a heme c. While iron is generally the central atom, this research will attempt to insert cobalt into the porphyrin ring of a heme c, creating cobalt-substituted heme proteins. While previously accomplished synthetically, this research will attempt to use E. coli to perform the cobalt insertion since past research has shown the process of cobalt insertion in the laboratory to be complicated and time consuming. These new proteins have applications in chemical catalysis and biofuel cells. The goal is to determine how much cobalt can be added to the E. coli under different conditions without killing the cells, such that cobalt insertion into porphyrin can be catalyzed by ferrochelatase. Next, the enzyme HCCS will need to attach the cobalt-substituted porphyrin, a non-native substrate, to the protein cytochrome c. We will be adding increasing amounts of cobalt to E. coli cells that express horse cytochrome c along with either the wild type HCCS or the mutant E138H that has been shown to relax the substrate specificity of HCCS. Additionally, we will be testing whether the overexpression of ferrochelatase affects the addition of cobalt. We expect the results to demonstrate greater expression levels of cobalt-substituted cytochrome c when co-expressed with mutant HCCS and ferrochelatase.

Expression of cobalt-substituted heme proteins in E. coli

Macromolecular Chemistry: The Second Century

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ACS Spring 2020 National Meeting & Expo

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.