Determining the Redox Potential of Heme, Embedded in Bacterioferritin Protein Shell, by Spectrophotometry: An Alternative Approach to Electrochemistry

Abstract

Iron homeostasis is critical to both host and pathogen, where ferritins (heme and non-heme binding) play a vital role. These protein nanocages sequester excess toxic, free iron and store them as iron biominerals. While most of the organisms synthesize only non-heme ferritin, bacteria such as Mycobacterium tuberculosis, the causative agent of tuberculosis, expresses both heme and non-heme binding ferritins. However, the exact role of heme remains an enigma despite our recent finding i.e., increasing heme content increases reductive iron release, suggesting its possible role as an electron mediator. Therefore, for understanding and regulating the electron transfer processes, determining the reduction potential (E1/2 ) of heme becomes important. In this regard, various electrode based analytical methods were found to be unsuccessful, as protein cage prevents heme-electrode contact. Here, we report a spectrophotometric method which not only maintains anaerobic conditions but facilitates heme reduction to determine the E1/2 value. These findings may not only help to understand the role of heme but also will guide to manipulate the heme environment for controlling microbial growth and developing bacterioferritin cage towards, new, non-natural functions