Two Routes for Extracellular Electron Transfer in Enterococcus faecalis

Abstract

Enterococcus faecalis cells are known to have ferric reductase activity and the ability to transfer electrons generated in metabolism to the external environment. We have isolated mutants defective in ferric reductase activity and studied their electron transfer properties to electrodes mediated by ferric ions and an osmium complex-modified redox polymer (OsRP). Electron transfer mediated with ferric ions and ferric reductase activity were both found to be dependent on the membrane-associated Ndh3 and EetA proteins, consistent with findings in Listeria monocytogenes In contrast, electron transfer mediated with OsRP was independent of these two proteins. Quinone in the cell membrane was required for the electron transfer with both mediators. The combined results demonstrate that extracellular electron transfer from reduced quinone to ferric ions and to OsRP occurs via different routes in the cell envelope of E. faecalisIMPORTANCE The transfer of reducing power in the form of electrons, generated in the catabolism of nutrients, from a bacterium to an extracellular acceptor appears to be common in nature. The electron acceptor can be another cell or abiotic material. Such extracellular electron transfer contributes to syntrophic metabolism and is of wide environmental, industrial, and medical importance. Electron transfer between microorganisms and electrodes is fundamental in microbial fuel cells for energy production and for electricity-driven synthesis of chemical compounds in cells. In contrast to the much-studied extracellular electron transfer mediated by cell surface exposed cytochromes, little is known about components and mechanisms for such electron transfer in organisms without these cytochromes and in Gram-positive bacteria such as E. faecalis, which is a commensal gut lactic acid bacterium and opportunistic pathogen.

Keywords: EetA; Enterococcus faecalis; PplA; extracellular electron transfer; ferric reductase; type 2 NADH dehydrogenase.

FIG 1

Map of the ndh3 gene region in the chromosome of E. faecalis. Transposon insertion positions in the five ferric reductase mutants isolated in this work are indicated with vertical arrows. For more information see Table 1.

FIG 2

Ferric reductase activity of E. faecalis parental (OG1RF) and mutant strains grown without heme. The strains are described in Table 1. (A) Activity with ammonium ferric sulfate. (B) Activity with potassium ferricyanide. Error bars show the standard error of the mean (SEM) based on more than four measurements with at least two biological replicates.

FIG 3

Electrochemical communication between heme-free E. faecalis parental (OG1RF) and mutant strains and graphite electrodes in the presence of various d-glucose concentrations. The strains are described in Table 1. (A and B) Current density responses with cells immobilized on OsRP-coated graphite electrodes. (C and D) Results for the cells immobilized on graphite electrodes and with ferricyanide (Fe³⁺) as the redox mediator. The electrochemical behavior of the mutant strains presented in panel C is shown in Fig. S2 with an expanded current density scale.