Adaptive Synthesis of a Rough Lipopolysaccharide in Geobacter sulfurreducens for Metal Reduction and Detoxification

Abstract

The ability of some metal-reducing bacteria to produce a rough (no O-antigen) lipopolysaccharide (LPS) could facilitate surface interactions with minerals and metal reduction. Consistent with this, the laboratory model metal reducer Geobacter sulfurreducens PCA produced two rough LPS isoforms (with or without a terminal methyl-quinovosamine sugar) when growing with the soluble electron acceptor, fumarate, but only expressed the shorter and more hydrophilic variant when reducing iron oxides. We reconstructed from genomic data conserved pathways for the synthesis of the rough LPS and generated heptosyltransferase mutants with partial (Δ rfaQ ) and complete (Δ rfaC ) truncations in the core oligosaccharide. The stepwise removal of the LPS core sugars reduced the hydrophilicity of the cell and increased outer membrane vesiculation. These changes in outer membrane charge and remodeling did not substantially impact planktonic growth but disrupted the developmental stages and structure of electroactive biofilms. Furthermore, the mutants assembled conductive pili for the extracellular mineralization of the toxic uranyl cation, yet were unable to prevent the permeation and mineralization of the radionuclide in the cell envelope. Hence, not only does the rough LPS promote cell-cell and cell-mineral interactions critical to biofilm formation and metal respiration, but it also functions as a permeability barrier to toxic metal cations. In doing so, the rough LPS maximizes the extracellular reduction of soluble and insoluble metals and preserves cell envelope functions critical to the environmental survival of Geobacter bacteria in metal rich environments and their performance in bioremediation and bioenergy applications. Importance Some metal-reducing bacteria produce a lipopolysaccharide (LPS) without the repeating sugars (O-antigen) that decorate the surface of most Gram-negative bacteria, but the biological significance of this adaptive feature has never been investigated. Using the model representative Geobacter sulfurreducens strain PCA and mutants carrying stepwise truncations in the LPS core sugars, we demonstrate the importance of the rough LPS in the control of cell surface chemistry during the respiration of iron minerals and the formation of electroactive biofilms. Importantly, we describe hitherto overlooked roles for the rough LPS in metal sequestration and outer membrane vesiculation that are critical for the extracellular reduction and detoxification of toxic metals and radionuclides. These results are of interest for the optimization of bioremediation schemes and electricity-harvesting platforms using these bacteria.

MliR, a novel MerR-like regulator of iron homeostasis, impacts metabolism, membrane remodeling, and cell adhesion in the marine Bacteroidetes Bizionia argentinensis

“…In line with this notion, missense mutations of kdsA gene have been linked to instability of the outer membrane, defects in FtsZ-ring formation and filamentous morphology in E. coli (Fujishima et al, 2002). In addition, several studies have addressed the relevance on LPS structure in modulating OM charge and facilitating cell attachment to surfaces, biofilm formation and metal uptake (Raetz et al, 2007;Clifton et al, 2016;Clark et al, 2021).…”

Electrochemical Characterization of the Periplasmic PpcA c‐Cytochrome of Geobacter sulfurreducens Reveals Its Affinity for Uranium

“…[8] Under such selective pressure, Geobacter cells have evolved detoxification mechanisms such as the release of outer membrane vesicles with the periplasmic minerals [8] and LPS-bound U VI . [14] Importantly, the cells assemble conductive pili to reductively precipitate the uranyl cation extracellularly. [8] Given the pleiotropic effects of many cytochrome [19] and pili [8,51] mutations on ET functions, the PpcA biomimetic interface may be enhanced with additional redox components of the Geobacter respiratory machinery to extend the kinetic and thermodynamic studies and provide additional insights into the bacterial mechanisms for metal reduction.…”

“…[13] In addition to the pili, the cells anchor on the outer membrane a rough (lacking the terminal repeating-sugar moiety or O-antigen) lipopolysaccharide (LPS) that exposes negatively charged ligands for the sequestration of the uranyl cation. [14] Despite the adaptive strategies that Geobacter cells employ to immobilize the uranyl cation extracellularly, some U traverses the outer membrane and is rapidly mineralized in the periplasmic space. [8,13] X-ray absorption spectroscopy analyses confirmed the reductive precipitation of the uranyl cation (provided as uranyl acetate) to a mononuclear phase of U IV in the periplasm.…”

“…Moreover, the extent of U reduction correlated linearly with the piliation levels [8] but not with the outer membrane c ‐cytochrome content [13] . In addition to the pili, the cells anchor on the outer membrane a rough (lacking the terminal repeating‐sugar moiety or O‐antigen) lipopolysaccharide (LPS) that exposes negatively charged ligands for the sequestration of the uranyl cation [14] …”

Molecular Mechanisms Underlying Bacterial Uranium Resistance

“…This could indicate that the extracellular pili function as a protective mechanism to avoid excessive precipitation in the periplasm, thereby minimizing cytotoxic effects. Furthermore, the metal-chelating properties of rough lipopolysaccharides could complement the extracellular pili by preventing uranium crossing the outer membrane, thereby creating a barrier to maximize extracellular reduction ( Clark et al, 2021 ). However, detailed knowledge about the mechanism is currently lacking.…”