Modular engineering to increase intracellular NAD(H/+) promotes rate of extracellular electron transfer of Shewanella oneidensis

Feng Li,Chen-Guang Liu,Yuan-Xiu Li,Liang Shi,Ying-Xiu Cao,Lei Wang,Hao Song

doi:10.1038/s41467-018-05995-8

Abstract

The slow rate of extracellular electron transfer (EET) of electroactive microorganisms remains a primary bottleneck that restricts the practical applications of bioelectrochemical systems. Intracellular NAD(H/+) (i.e., the total level of NADH and NAD+) is a crucial source of the intracellular electron pool from which intracellular electrons are transferred to extracellular electron acceptors via EET pathways. However, how the total level of intracellular NAD(H/+) impacts the EET rate in Shewanella oneidensis has not been established. Here, we use a modular synthetic biology strategy to redirect metabolic flux towards NAD+ biosynthesis via three modules: de novo, salvage, and universal biosynthesis modules in S. oneidensis MR-1. The results demonstrate that an increase in intracellular NAD(H/+) results in the transfer of more electrons from the increased oxidation of the electron donor to the EET pathways of S. oneidensis, thereby enhancing intracellular electron flux and the EET rate.

Highlights

The slow rate of extracellular electron transfer (EET) of electroactive microorganisms remains a primary bottleneck that restricts the practical applications of bioelectrochemical systems
Electrons derived from the oxidation of carbon sources and electron donor by S. oneidensis MR-1 flow through an intracellular electron transfer pathway, namely, from NADH through intracellular menaquinol pools and a metal-reducing (Mtr) pathway, including a cytoplasmic membrane c-type cytochrome (c-Cyt) CymA, the periplasmic c-Cyts STC and FccA, and the outer membrane “porin-cytochrome” (OmcA-MtrCAB) to the outer membrane[33,34], which subsequently transfer to the extracellular electron acceptors via the contactedbased EET pathway based on c-Cyts[35] and soluble electron shuttle-mediated EET pathways based on flavins as electron shuttles[31,36] or bound cofactors for MtrC and OmcA37,38
Our results suggest that the total NAD(H/+) is a crucial intracellular electron pool, and an increase in this pool could result in the transfer of more electrons from increased oxidation of the electron donor to the EET pathways of S. oneidensis, thereby enhancing intracellular electron flux and the EET rate

Summary

Introduction

The slow rate of extracellular electron transfer (EET) of electroactive microorganisms remains a primary bottleneck that restricts the practical applications of bioelectrochemical systems. Electrons derived from the oxidation of carbon sources and electron donor (i.e., lactate) by S. oneidensis MR-1 flow through an intracellular electron transfer pathway, namely, from NADH (a typical carrier of intracellular electrons) through intracellular menaquinol pools and a metal-reducing (Mtr) pathway, including a cytoplasmic membrane c-type cytochrome (c-Cyt) CymA, the periplasmic c-Cyts STC and FccA, and the outer membrane “porin-cytochrome” (OmcA-MtrCAB) to the outer membrane[33,34], which subsequently transfer to the extracellular electron acceptors (e.g., metal oxides or electrodes) via the contactedbased EET pathway based on c-Cyts[35] and soluble electron shuttle-mediated EET pathways based on flavins as electron shuttles[31,36] or bound cofactors for MtrC and OmcA37,38 Based on these two underlying EET mechanisms, a number of synthetic biology strategies have been developed to enhance the rate of EET in S. oneidensis. Our study shows that synthetic biology strategies to increase the intracellular NAD(H/+) level are of great value in increasing the EET rate of exoelectrogens

Methods

Results

Conclusion