(582aw) 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).