Computational and Experimental Analysis of Redundancy in the Central Metabolism of Geobacter sulfurreducens

Daniel Segura,Radhakrishnan Mahadevan,Derek R Lovley,Katy Juárez,Roy Kishony

doi:10.1371/journal.pcbi.0040036

Abstract

Previous model-based analysis of the metabolic network of Geobacter sulfurreducens suggested the existence of several redundant pathways. Here, we identified eight sets of redundant pathways that included redundancy for the assimilation of acetate, and for the conversion of pyruvate into acetyl-CoA. These equivalent pathways and two other sub-optimal pathways were studied using 5 single-gene deletion mutants in those pathways for the evaluation of the predictive capacity of the model. The growth phenotypes of these mutants were studied under 12 different conditions of electron donor and acceptor availability. The comparison of the model predictions with the resulting experimental phenotypes indicated that pyruvate ferredoxin oxidoreductase is the only activity able to convert pyruvate into acetyl-CoA. However, the results and the modeling showed that the two acetate activation pathways present are not only active, but needed due to the additional role of the acetyl-CoA transferase in the TCA cycle, probably reflecting the adaptation of these bacteria to acetate utilization. In other cases, the data reconciliation suggested additional capacity constraints that were confirmed with biochemical assays. The results demonstrate the need to experimentally verify the activity of key enzymes when developing in silico models of microbial physiology based on sequence-based reconstruction of metabolic networks.

Highlights

Geobacter species are of interest because of their natural role in carbon and mineral cycling, their ability to remediate organic and metal contaminants in the subsurface, and their capacity to harvest electricity from waste organic matter [1,2,3]
An understanding of acetate metabolism in Geobacter species is required because acetate, secreted by fermenting organisms, is the dominant electron donor for Geobacteraceae in soils and sediments [15], and because recent studies have shown that the addition of acetate to uranium-contaminated aquifers can stimulate in situ bioremediation of uranium contamination by Geobacter species [16,17]
Geobacter sulfurreducens is a member of the Geobacteraceae family of micro-organisms that breathe metals and have a unique mode of metabolism

Summary

Introduction

Geobacter species are of interest because of their natural role in carbon and mineral cycling, their ability to remediate organic and metal contaminants in the subsurface, and their capacity to harvest electricity from waste organic matter [1,2,3]. G. sulfurreducens can use either acetate or hydrogen as the sole electron donors for Fe(III) reduction, and fumarate or malate can be used as terminal electron acceptors [4]. To better understand the physiology of G. sulfurreducens, a constraint-based genome-scale metabolic model was constructed and used to investigate the unique physiology associated with the reduction of extracellular electron acceptors, such as Fe(III) [21]. The genome-scale model enabled the assessment of the impact of global proton balance during Fe(III) reduction on biomass and energy yields, and successfully predicted the lower biomass yields observed during the growth of a mutant in which the fumarate reductase had been deleted [22].

Author Summary

Findings

Materials and Methods