Date of Award
Doctor of Philosophy (PhD)
Several anoxygenic phototrophs grow by utilizing soluble iron or insoluble mixed-valence iron minerals (such as rust) as electron donors to fix carbon dioxide using light energy, a process called photoferrotrophy. Photoferrotrophs can also use electron donors such as poised electrodes that serve as proxies for rust via phototrophic extracellular electron uptake (EEU). Despite the recognition that these two related microbial processes contribute to various biogeochemical cycles such as iron and carbon, the electron uptake mechanisms underlying photoferrotrophy and phototrophic EEU are poorly understood. To address the key knowledge gaps in our understanding of these microbial metabolisms, here we characterized Rhodopseudomonas palustris TIE-1 as a model photoferrotrophic bacterium via a combinatorial approach using genetics, biochemistry, microfluidics, and bioelectrochemistry. This study demonstrates that photoferrotrophs such as R. palustris TIE-1 produce a porin-cytochrome complex at its surface, which acts as an electron conduit to harvest electrons across the outer membrane from both soluble iron and poised electrodes. The electron conduit consists of a single periplasmic decaheme cytochrome c (PioA) and an outer membrane porin (PioB). To the best of our knowledge, this is the first report of an EEU mechanism in a photoautotrophic microbe. This mechanism is distinct from those used by non-phototrophic bacteria because the PioAB electron conduit contains no apparent extracellular cytochrome c component. This study shows that PioA undergoes postsecretory proteolysis of its N-terminus to produce a shorter heme-attached PioA (holo-PioAC, where PioAC represents the C-terminus of PioA). The holo-PioAC is an iron oxidoreductase that exists freely in the periplasm and forms a membrane-associated complex with PioB. The extended N-terminal peptide controls heme attachment, and its processing is required to produce wild-type levels of holo-PioAC and holo-PioACB complex. The postsecretory proteolysis of the N-terminal extension of PioA-like homologs to produce a functional holo-PioAC and the formation of holo-PioACB electron conduit are conserved in phototrophs harboring homologous proteins such as Rhodomicrobium vannielii and Rhodomicrobium udaipurense. The presence of PioAB in these organisms correlates with their ability to perform photoferrotrophy and phototrophic EEU. More importantly, findings from this study are consistent with the idea that the Gram-negative phototrophs, oxidize soluble iron extracellularly because the product of this process is insoluble iron, which is lethal if accumulated in the periplasm. Together, our results suggest that freshwater photoferrotrophic bacteria are also capable of EEU. However, the EEU capability and the underlying electron transfer mechanisms of marine photoferrotrophs is unknown. To fill up this knowledge gap, here we also characterized photoferrotrophy and phototrophic EEU in a newly isolated marine phototrophic bacterium, Rhodovulum sulfidophilum AB26 (AB26). We show that AB26 can perform both photoferrotrophy and phototrophic EEU. AB26 uses a range of potentials that mimic the midpoint potential of substances that are abundant in marine settings such as mixed-valence iron, iron-sulfur, and elemental sulfur-containing minerals. Our study indicates that the electrons taken up during phototrophic EEU enter the photosynthetic electron transport chain, and phototrophic EEU results in the upregulation of carbon fixation and storage pathways. This is consistent with the idea that electron uptake during phototrophic EEU is linked to carbon fixation in AB26. Based on comparative genomics, transcriptomics, and proteomics analyses, we suggest that AB26 uses an extracellular electron transfer mechanism that is different from TIE-1. In this study, we also developed a genetic system de novo in AB26 for future genetic studies focusing on identifying the EEU pathway and its molecular mechanism. Overall, these studies suggest that photoferrotrophy and phototrophic EEU are widespread processes. EEU could enhance microbial survival and support primary productivity in anoxygenic environments deficient in soluble electron donors but abundant in redox-active minerals such as those containing iron. Via their growth and activity, phototrophic EEU-capable microbes play an important role in the global biogeochemical cycling of elements like iron and carbon on Earth. In addition, photoferrotrophy is considered a potential metabolism responsible for the deposition of the Archean banded iron formations (BIFs). Findings from this study will be helpful to our understanding of metabolic evolution, microbial ecology, and Earth history. Microbes capable of phototrophic EEU are good candidates for microbial electrosynthesis, a process in which microbes use electricity to produce biocommodities from carbon dioxide. Findings from this research will aid in improving such biotechnological applications and empower synthetic biology.
Chair and Committee
Robert Kranz, Joseph Jez, Petra Levin, Meredith Jackrel,
Gupta, Dinesh, "Understanding the Molecular Mechanisms of Photoferrotrophy and Phototrophic Extracellular Electron Uptake" (2021). Arts & Sciences Electronic Theses and Dissertations. 2320.