Engineering Rhodobacter Sphaeroides for Direct Electron Transfer By Heterogenous Expression of Redox-Active Components From Electrogenic Species

Engineering Rhodobacter  Sphaeroides for direct electron transfer by heterogenous expression of redox-active components from electrogenic species 

Danhui Cheng^1,2, Wanyang Wang^1,2, King L Chow^1,2,4, and I-Ming Hsing^1,2,3*

 

Bioengineering Graduate Program¹, Division of Biomedical Engineering², Department of Chemical and Biomolecular Engineering³, Division of Life Science⁴, the Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong, SAR China

(* Corresponding Author)  Fax: 852-2358-0054, E-mail : kehsing@ust.hk

Direct electron transfer between photosynthetic bacteria and electrode surface has been pursued in the study of bioelectricity generation through a solar powered microbial fuel cell, with the purpose to eliminate the usage of redox mediators and enhance the efficiency of power generation. In nature, a few species, including Shewanella oneidensis and Geobacter sulfurreducens are able to ‘respire' on insoluble iron oxide through direct electron flow from cytoplasmic oxidative reactions to metal oxide/electrode surface. This feature attributes to special electron transfer conduit composed of cytochromes and structural proteins. A previous study has shown that expressing key components in the electron conduit of Shewanella can convert E.coli into microorganism vehicle capable of direct electron transfer. [1]

In this presentation, we aim to construct a direct electron transfer pathway in a photosynthetic purple non-sulfur bacterial species, Rhodobacter sphaeroides via heterologous expression of MtrCAB complex from Shewanella oneidensis. MtrCAB consists of periplasmic decaheme cytochrome MtrA, outer membrane β-barrel protein MtrB, and outer membrane decaheme cytochrome MtrC. Through the interaction of MtrA with cytoplasmic membrane components, electron flow can be channeled from cytoplasmic quinol pool to the periplasmic region. MtrA can then interact with MtrC through MtrB and direct the electron to outer membrane. The electron can be ultimately delivered to an external metal oxide/electrode surface via the direct contact between the surface and MtrC.

Figure. Engineering Rhodobacter for direct electron transfer by expression of MtrCAB complex for application in a solar powered microbial fuel cell.

In our preliminary study, decaheme cytochrome MtrA was first expressed in Rhodobacter. Using the native signal peptide from Rhodobacter cytochrome c₂, we successfully localize MtrA in the periplasmic region. Both TMB-based heme staining and absorption spectrometry have shown that the heme group is correctly incorporated in MtrA and confers its redox activity. The MtrA elevates the reduction rate of soluble ferric iron in Rhodobacter when a highly reduced substrate, butyrate, is provided (i.e., when the cytoplasmic quinol pool of Rhodobacter is over-reduced). The preliminary results implicate feasibility of redirecting electrons from cytoplasmic oxidative reactions to the extracellular electron pathway in Rhodobacter through this synthetic electron conduit.

[1] Jensen, H. M., Albers, A. E., Malley, K. R., Londer, Y. Y., Cohen, B. E., Helms, B. a, Weigele, P., et al. Engineering of a synthetic electron conduit in living cells. Proceedings of the National Academy of Sciences of the United States of America, 107(45), 19213–8(2010).