Sulfur is one of the most important and versatile elements in biology and forms the basis of energy metabolism in many prokaryotes. Its extensive large-scale redox transformations drive the global biogeochemical sulfur cycle and profoundly influence environmental chemistry. In dissimilatory sulfur oxidizers, the oxidation of reduced sulfur compounds is directly coupled to energy conservation via photosynthetic or respiratory electron transport chains. Sulfur-oxidizing bacteria and archaea are remarkably diverse, reflecting both the wide range of habitats they inhabit and the variety of metabolic pathways through which sulfur compounds are exploited as electron donors. Over the past decades, these pathways have been examined in depth using a spectrum of molecular genetic, biochemical, and omics-based approaches, primarily in model organisms amenable to genetic manipulation. Particular emphasis has been placed on thiosulfate oxidation mediated by the Sox multienzyme system and thiosulfate dehydrogenases, as well as on cytoplasmic sulfane sulfur oxidation involving dissimilatory sulfite reductase and the sulfur-oxidizing heterodisulfide reductase-like sHdr complex. Recent advances have highlighted the central role of specific lipoate-binding proteins that are essential for efficient sHdr-dependent sulfur oxidation. This review provides an overview of current knowledge on prokaryotic sulfur oxidation pathways. In addition, emerging insights into the complex regulatory networks operating in facultative sulfur oxidizers are provided, with particular attention to how these organisms coordinate sulfur metabolism with changing environmental and energetic conditions.
Bacteria obtain nutrients and other essential factors from their immediate surrounding. For bacteria that reside in a specialized or limited niche range, their metabolism reflects adaptation to that environment(s). Bordetella pertussis and B. parapertussis have evolved from a B. bronchiseptica-like ancestor that survives within hosts and in the environment to become restricted to a single niche, the human respiratory tract. This restriction has shaped their metabolism. Here, the multifaceted role of nicotinic acid in Bordetella biology is reviewed. It serves as a precursor for the synthesis of NAD, for which these Bordetella are auxotrophic, but also induces profound changes in gene expression profiles in a concentration-dependent manner. While these Bordetella display very high levels of conservation in many aspects, they differ in their utilization of nicotinic acid. Understanding the consequences of this difference may shed light on host-pathogen interactions of these niche specialized species.
Glycyl radical enzymes (GRE) use radical-based biochemistry to catalyze the cleavage or formation of the carbon-carbon and carbon-heteroatom bonds found in organic molecules. These evolutionarily ancient enzymes are highly oxygen-sensitive but widespread in obligate as well as facultative anaerobes. When synthesized, GRE lack activity. The radical must be installed through the action of an activating enzyme (AE) that belongs to the radical S-adenosylmethionine superfamily. Typically, each GRE has a dedicated GRE-AE. Hydrogen atom abstraction from a highly conserved glycine residue located within the GRE's C-terminal domain introduces the radical into its storage location. Upon substrate binding, the radical is transferred to a conserved cysteine residue within the active site, transiently generating a thiyl radical species and initiating catalysis. There are currently five biochemical classes of GRE. Nevertheless, these GRE classes share a common mechanism of glycyl-radical storage and thiyl radical-mediated generation of a substrate radical. GRE exhibit similar tertiary structures and conserved active sites, but otherwise show limited primary structural conservation. These features are consistent with them diverging from an ancient common ancestor. Recent genome mining analyses, combined with structural and biochemical studies, are revealing the ubiquity of GRE, especially in uncultured gut microbiota and in association with disease states. Moreover, GRE are predominant in environmental microbiota associated with anaerobic degradation of hydrocarbons. GRE now have mainly catabolic roles in fermentative heterotrophs, but their universal involvement in anaerobic DNA synthesis and in other chemically challenging reactions suggests earlier in evolution biosynthesis of organic molecules might have been their main function.
Iron-sulfur (Fe-S) clusters are essential cofactors that support a wide range of metabolic and regulatory processes across all domains of life. The assembly and distribution of these chemically labile cofactors require tightly coordinated biosynthetic and trafficking systems that respond dynamically to cellular iron availability and redox conditions. Monothiol CGFS-type glutaredoxins (Grxs) have emerged as central components of these networks. Once considered primarily thiol-disulfide oxidoreductases, these proteins are now recognized as versatile Fe-S cluster carriers that coordinate and exchange [2Fe-2S] clusters through glutathione (GSH)-dependent mechanisms. This review synthesizes current understanding of monothiol Grxs across bacteria, fungi, and protists, highlighting both conserved biochemical functions and lineage-specific adaptations. In all systems, CGFS Grxs function as intermediates in Fe-S cluster trafficking pathways, facilitating the transfer of clusters from assembly machineries to downstream targets. In fungi, these proteins have been further co-opted into regulatory circuits, where Grxs alone or in partnership with BolA proteins directly couple mitochondrial Fe-S cluster biogenesis to nuclear transcriptional control of iron homeostasis. In contrast, bacterial and protist systems exhibit more indirect or emerging regulatory roles, often integrating Fe-S metabolism with broader redox and stress-response networks. We propose a unifying model in which monothiol Grxs act as dynamic Fe-S "rheostats" that sense and redistribute labile clusters in response to cellular conditions, thereby linking iron metabolism to physiological adaptation. Understanding how this conserved molecular framework is differentially deployed across organisms provides new insight into microbial iron homeostasis and reveals potential targets for therapeutic intervention in pathogenic systems.
Electronic waste (E-waste) is one of the fastest growing waste streams worldwide, posing serious challenges for environmental and human health, while representing a rich secondary source of valuable metals. Among these technology-critical metals (TCMs) are those, which are essential for renewable energy systems, electronics and low carbon technologies, yet face increasingly disrupted supply chains. This chapter explores the role of E-waste as an urban mine within a circular economy framework and highlights the emerging bio-based strategies for sustainable metal recovery, with particular emphasis on bacterial systems. Bacteria employ versatile mechanisms such as biosorption, bioaccumulation, bioleaching and biomineralization to capture, dissolve and transform metals, offering a green alternative to conventional physical and chemical methods for metal recovery. Beyond recovery, bacteria can upcycle the recovered metals into value added nanoparticles with diverse biomedical, environmental and catalytic applications. Advanced omics approaches including transcriptomics, metabolomics and proteomics provide key insights for the rational design of synthetic biology strategies aimed to enhance metal recovery and enable controlled, efficient synthesis of metal nanoparticles with tailored properties. Overall, the chapter combines E-waste management with microbial biotechnology and nanotechnology presenting a sustainable way to secure technology-critical metals, reducing environmental impact and promoting the transition to a green circular economy.
Fifty years ago, the discovery of superoxide dismutase revealed that reactive oxygen species (ROS) comprise an unavoidable stress for organisms that dwell in oxic habitats. Since then, workers have gradually pieced together the details. Superoxide and hydrogen peroxide are continuously generated in aerobic cells when oxygen collides with the exposed cofactors of redox enzymes. The rate of electron spillage varies from enzyme to enzyme, and the primary ROS sources have not yet been identified, in any organism. Superoxide and hydrogen peroxide can disrupt key metabolic pathwfays by specifically damaging the iron-sulfur clusters and ferrous cofactors of enzymes. Hydrogen peroxide also threatens DNA, by generating hydroxyl radicals through its oxidation of loose iron. Oxygen-tolerant bacteria successfully suppress these problems by synthesizing high titers of scavenging enzymes. However, hydrogen peroxide can also be formed in the environment, and when it penetrates cells, it can overwhelm basal defenses. Accordingly, it seems universally true that microbes possess specialized transcription factors that detect any rise in peroxide levels and activate tactics to defend their enzymes and DNA. This review describes the chemistry of oxidative stress and the multilayered survival strategies of bacteria. Workers are now focused upon identifying real-world circumstances in which ROS stress is so severe that it becomes bacteriostatic or lethal. Of particular interest is the likelihood that biological warfare leverages ROS through the actions of redox-active antibiotics and cell-based immune responses.
Extracellular electron transfer (EET) may well have been the earliest form of microbial respiration, but it is also one of the most recently discovered. EET plays an important role in the biogeochemical cycling of carbon, metals, and nutrients; corrosion of metals; the conversion of organic wastes to methane; and the bioremediation of subsurface contaminants. A broad diversity of bacteria and archaea, inhabiting a wide range of environments, are capable of EET. Limited study of just a few model microbes has already revealed multiple divergent EET mechanisms for electron transfer to the outer cell surface. Although most EET studies have focused on microbes that form electrical contacts at their outer surface, intracellularly reduced electron shuttles may also be widespread. Effective growth with minerals like Fe(III) oxides as the electron acceptor also requires solubilization with chelators; redox-active electron shuttles to ferry electrons from the cell surface to electron acceptors; or the expression of outer-surface nanowires to extend the electronic reach of the cells. Functional studies on nanowires have been restricted to Geobacter species, which produce electrically conductive pili that are required for long-range EET, and cytochrome filaments. Promoting direct interspecies electron transfer to improve the conversion of organic wastes to methane is an emerging focus of applied EET research, as is the prevention of EET associated with microbial metal corrosion. Microbes' ability to exchange electrons with electrodes have inspired multiple bioelectrochemical technologies. Electroactive microbes are the sensing component of novel living electronic sensors. e-Pili have been incorporated in electronic devices with unique sensing, electricity generation, and neuromorphic functions. However, most EET-related applications are in early phases of development. Future research opportunities in diverse aspects of EET are highlighted throughout.
Polyhydroxyalkanoates (PHAs) are a diverse family of intracellular polyesters synthesized by bacteria and archaea across a wide array of metabolic and ecological contexts. Originally characterized as inert carbon and energy reserves, PHAs are now recognized as multifunctional biopolymers with dynamic roles in redox homeostasis, stress mitigation, and microbial community interactions. This review presents a comprehensive synthesis of the current understanding of PHA metabolism-from canonical biosynthetic enzymes and granule-associated structural proteins to diverse regulatory circuits and pathway variants. We highlight recent discoveries in anaerobic and facultative aerobic PHA producers, uncover emerging patterns in ecological function, and integrate novel computational approaches-including Hidden Markov Models, protein language models, and AI-guided enzyme design-for functional gene discovery. Special attention is given to the redox buffering capacity of PHA granules, their role in stress resilience, and their contribution to microbial fitness in fluctuating environments. Finally, we examine the techno-economic landscape of microbial PHA production, the application of PHA-derived materials in biomedical and industrial contexts, and the broader implications for sustainable biopolymer design. This review synthesizes recent advances in PHA biology, reframing these polymers from inert carbon and energy depots to active, dynamic participants in microbial physiology, integrating redox homeostasis, metabolic flux regulation, and adaptive stress responses.
The existence of steroid antibacterials has been studied since the golden age of antibiotics, although the literature on this topic is fragmented. To address this the current review seeks to bring these fragmented lines of enquiry together to assess the utility of steroidal compounds as antibacterials. In doing so this review will document the structural features, biosynthetic/synthetic origins, antibacterial toxicities, potential for fungal biotransformations, and catabolic pathways in the human host. Herein, we identify steroid antibacterials from fungal origins, synthetic steroids, and approved drugs that have the potential for future repurposing as antibacterials. Of particular note are the fusidane steroids of fungal origin, displaying low MIC values and being effective against a variety of Gram-positive and Gram-negative species. At present, fusidic acid is the only steroid that is prescribed as an antibacterial, despite many publications describing antibacterial activity in a range of well-known steroids. The impact of bacterial exposure to steroids naturally differs depending on the nature of the cell membranes, cell wall, and the ability of bacterial to import steroids for use as a carbon source, which is discussed herein. Also, this study explores the biotechnological potential of fungi for the production of antibacterial steroids, and how steroid drugs are metabolised in the human host. Together, this provides an assessment of the potential for steroids as future antibacterial drugs.
Transport is instrumental to all aspects of life. At the microbial level, transport mechanisms mediate physiology, nutrition and defence, but also impact clinical medicine, requiring consideration in drug mode of action through to target-cell efflux. Whereas efflux of toxic compounds is critical to survival of all cells, this evolutionarily conserved mechanism also mediates the medical threat of antibiotic resistance. Multidrug transporters are therefore a large and important family of proteins across evolution. Their function in membranes of the eukaryotic model microbe, Dictyostelium discoideum, are not well documented, despite exquisitely dissected genetics and mechanics for transport via membrane trafficking. MDR proteins in this amoeba are worthy of greater attention, however. The Dictyostelium genetic complement has proven relevance to many aspects of human disease pathology and pathogen interactions with host, whereas transporter roles have been little-explored in the organism's fascinating life cycle of chemotaxis, cooperation and cell differentiation. By documenting the MDR protein complement in D. discoideum, compiling the sparse reports of experimentally characterised transporters, and integrating RNAseq through the life cycle with transcriptomics co-expression data, this review aims to highlight questions and key areas of interest for the ABC, MFS, MATE, SMR/DMT, RND, AbgT, and PACE transporter groups. Whether as parts for synthetic biology application, natural product biosynthetic gene cluster identification, or for understanding resistance in pharmaceutical research, there are key applied areas of learning alongside discovering evolutionarily conserved roles in microbial physiology.
Paracoccus denitrificans is a long-established model organism for studies of methylotrophy, the use of one-carbon compounds as sources of energy and carbon. P. denitrificans can use methanol and methylamine as growth substrates, oxidizing both to formaldehyde in the periplasm. Formaldehyde is oxidized to formate and then to carbon dioxide, which is assimilated into biomass via the Calvin cycle. Genes required for the oxidation of methanol, methylamine, formaldehyde and formate are typically expressed only under methylotrophic conditions or during growth on multi-carbon substrates (such as choline) the catabolism of which generates formaldehyde as a product of demethylation reactions. In this article, we review the pathways of methylotrophic metabolism and the proteins involved, before focusing on mechanisms of gene regulation. P. denitrificans has genes encoding calcium- and lanthanide-dependent methanol dehydrogenases. In other methylotrophs, expression of these enzymes is subject to reciprocal regulation according to the presence or absence of lanthanide ions in growth media. This regulatory phenomenon is referred to as the 'lanthanide switch'. We propose a model for the mechanism of the lanthanide switch in P. denitrificans, which extrapolates from relevant information in other methylotrophs and is consistent with prior literature.
In global biogeochemical networks nitrogen transitions between a number of different oxidation states, from +5 to -3. The two most oxidized states are found in the nitrogen oxyanions nitrate (NO3-, +5) and nitrite (NO2-, +3). These two oxyanions form an electropositive redox couple, with a midpoint redox potential (pH7) of +430 mV, that enables them to serve as both electron acceptor (nitrate) and electron donor (nitrite) in a range of catabolic and anabolic processes. Several enzymatic systems have been identified that can inter-convert the two oxyanions and couple them to a range of electron transport pathways. Recent literature on nitrate reduction and nitrite oxidation by prokaryotes reveals a great number of meta "omics" studies identifying genes, transcripts or peptides functionally related to the nitrate / nitrite redox couple in a wide range of environments. To fully interpret such data in the context of the environment being studied requires a recognition of the different physiological functions the nitrate / nitrite redox couple is able to support. This, in turn, is related to the biochemical diversity of the enzymes that drive this reversible redox couple in nature. This review seeks to define bioenergetically the different enzymes involved in nitrate-nitrite inter-conversion and relate this to the diverse physiological activities that this redox couple supports.
Nitrous oxide is a major contributor towards greenhouse gas emissions from agriculture and is the most significant single cause of ozone depletion in the 21st Century. In this chapter, the microbial processes associated with the production and consumption of nitrous oxide are reviewed, with a focus on the role of NosZ in nitrous oxide removal. Recent developments have led to a recognition that two distinct clades of nosZ exist, and that diversity exists within and between the clades resulting in functional diversity of NosZ in the organisms that carry them. We point out areas where there are knowledge gaps, particularly a lack of exploration of the comparative biochemistry of NosZ from organisms beyond a few laboratory model species. We discuss the importance of considering how nitrous oxide is measured, and the ways in which factors such as evolutionary selection pressure, regulation, and biochemical organisation impact on the eventual activity of nitrous oxide reduction in biological ecological systems. This is followed by a set of perspectives on how we might apply our current and future knowledge to mitigate atmospheric nitrous oxide accumulation for global benefit.
Vitamins are indispensable cofactors that expand the chemical capabilities of enzymes beyond the inherent limitations of amino acid side chains. Among them, vitamin B₁₂ is particularly remarkable due to its exceptional structural complexity, the presence of a cobalt-centered corrin ring, and its exclusive biosynthetic origin in prokaryotes. This review explores the biosynthesis, transport, and biological significance of B₁₂, with an emphasis on the growing toolbox of synthetic analogues designed for research and therapeutic use. Recent advances in synthetic biology have enabled the complete heterologous expression of the aerobic B12 biosynthesis pathway in Escherichia coli, facilitating the high-yield production of biosynthetic intermediates and cobalt-free B12-precursors. These intermediates serve as platforms for the generation of metbalamins, metal-substituted cobalamin analogues incorporating rhodium, nickel, zinc, and other transition metals. In parallel, novel organo-antimetabolites and fluorescently labelled derivatives have been developed to probe B₁₂-dependent enzymes, trace vitamin transport in living systems, and selectively disrupt microbial or disease-linked metabolism. These synthetic analogues function as versatile tools for imaging, mechanistic dissection, and metabolic inhibition and more specifically in the case of molecules that counteract the physiological effects of vitamin B12 in animal systems hold potential as antivitamins B12. Collectively, they offer powerful new approaches to study nutrient trafficking, engineer cofactor interactions, and develop targeted antimicrobial or anticancer strategies. The review concludes by discussing future directions in applying engineering biology and chemical synthesis to further diversify and exploit the functional potential of the cobalamin scaffold.
Stuart Ferguson made important contributions to our understanding of many aspects of bioenergetics, including the operation of the ATP synthase, all of the steps involved in denitification and the mechanism of cytochrome c biogenesis. In this article, we outline his career and describe the development of his research, highlighting his insights into the role of the bacterial periplasm in electron transport reactions and the diversity of covalent attachment mechanisms of heme to apo-cytochrome c.
Disulfide bonds are covalent linkages connecting two cysteine residues. When formed within the same polypeptide, they assist protein folding and enhance protein stability. In principle, disulfide formation could be facilitated by ubiquitous small-molecule oxidants, like oxygen. Instead, it is catalyzed by dedicated oxidative protein folding pathways throughout the tree of life. In bacteria, disulfides are abundant outside the cytoplasm, whereby chemical and mechanical stresses take their toll on protein molecules. The Disulfide Bond Formation (DSB) system in Escherichia coli K-12 has served as the paradigm for bacterial disulfide bond formation and has been, largely, considered a proteome housekeeper. In this article we discuss the central role of the DSB system for protein homeostasis, the unprecedented diversity of DSB proteins across the bacterial phylogeny, and their emerging roles in infectious disease. We also propose that beyond the known uses of DSB components in biotechnology, the DSB system offers promising avenues for the development of next-generation strategies against challenging bacterial pathogens.
This review focuses on some of the persisting misconceptions and even errors in the literature of bacterial denitrification and the respiratory reduction of nitrate to ammonia. Both processes were traditionally investigated using pure culture laboratory techniques and substrate concentrations in the high micromolar or millimolar range. These concentrations are 1000-fold higher than those found in the nanomolar natural environments in which bacterial metabolism continues to evolve. Many of the enzymes involved in anaerobic nitrate reduction are metalloproteins that are easily inactivated by exposure to reactive oxygen and reactive nitrogen species. However, the metal centers of some of these proteins retain the ability to catalyze chemical reactions irrelevant to their physiological function. The review highlights some of the errors and misconceptions persisting in the literature, especially in the context of sensing, production and reduction of nitric oxide. It challenges many statements about physiological relevance. It demonstrates how knowledge of mechanisms that regulate gene transcription and mRNA translation provide clues to enzyme function. Four criteria are proposed to judge whether a protein-dependent reaction is physiologically relevant. They include whether (i) the protein is present in the correct cellular location; (ii) its synthesis is regulated in response to, or in preparation for, its proposed role; (iii) the catalytic efficiency is adequate to fulfil the need; and (iv) alternative enzymes are available that better meet the first three criteria. How errors become embedded in the literature, perpetuated and reinforced by annotation errors in genome databases are highlighted.
Lactate is a key metabolite that is used as a carbon and energy source. It can also be generated as a metabolic end product, through reduction of pyruvate. Bacterial enzymes involved in lactate generation are classified as NAD+-dependent lactate dehydrogenases and are generally involved in production of lactate during fermentation, while NAD+-independent lactate dehydrogenases are involved in oxidation of lactate that is linked to reduction of quinone in respiratory or photosynthetic electron transport pathways, or in anaerobic lactate oxidation linked to electron bifurcation during heterotrophic growth. Enzymes specific for D-lactate, L-lactate or both stereoisomers exist and interconversion of D- and L- stereoisomers is catalyzed by a lactate racemase. Expression of operons encoding enzymes and transporters involved in lactate metabolism is regulated in several ways that can include sensing of the presence of L- or D- lactate by transcriptional regulators, control of gene expression through global regulators of carbon metabolism and regulators that respond to iron availability. Sensing of lactate also appears to be an important cue for changes in cell physiology and behavior and in some bacteria it has been shown to influence biofilm formation. Lactate plays a key role in the maintenance of human microbiomes in different niches and dysbiosis is often a result of an imbalance between lactate production and lactate consumption, which is linked to certain pathologies. Lactate is also an important carbon source for some bacterial pathogens and L-lactate has been shown to play a role in the pathogenesis in animal models of infection. Additionally, L-lactate produced by macrophages, neutrophils and epithelial cells may provide an important carbon source of the survival and growth of intracellular pathogens. Understanding of lactate metabolism at the biochemical, cellular and organismal/community level is of major importance in understanding and management of health and disease and in understanding environmental processes.
Photosynthesis by (bacterio)chlorophyll-producing organisms ("chlorophototrophy") sustains virtually all life on Earth, providing the biosphere with food and energy. The oxygenic process carried out by plants, algae and cyanobacteria also generates the oxygen we breathe, and ancient cyanobacteria were responsible for oxygenating the atmosphere, creating the conditions that allowed the evolution of complex life. Cyanobacteria were also the endosymbiotic progenitors of chloroplasts, play major roles in biogeochemical cycles and as primary producers in aquatic ecosystems, and act as genetically tractable model organisms for studying oxygenic photosynthesis. In addition to the Cyanobacteriota, eight other bacterial phyla, namely Proteobacteria/Pseudomonadota, Chlorobiota, Chloroflexota, Bacillota, Acidobacteriota, Gemmatimonadota, Vulcanimicrobiota and Myxococcota contain at least one putative chlorophototrophic species, all of which perform a variant of anoxygenic photosynthesis, which does not yield oxygen as a by-product. These chlorophototrophic organisms display incredible diversity in the habitats that they colonise, and in their biochemistry, physiology and metabolism, with variation in the light-harvesting complexes and pigments they produce to utilise solar energy. Whilst some are very well understood, such as the proteobacterial 'purple bacteria', others have only been identified in the last few years and therefore relatively little is known about them - especially those that have not yet been isolated and cultured. In this chapter, we aim to summarise and compare the photosynthetic physiology and central metabolic processes of chlorophototrophic members from the nine phyla in which they are found, giving both a short historical perspective and highlighting gaps in our understanding.
We begin by discussing some historical ideas about the natural dynamics of living organisms and their complex states, from the very transient to the very persistent. We classify ultradian rhythms as being more important than most oscillatory behaviour in their distinctive properties. Then we summarise rhythmicity in three yeasts and five representative protist species. Recent discoveries about the single-celled photosynthetic alga Chlamydomonas reinhardii are then discussed as revealed by computer-controlled semi-continuous monitoring: automatic periodic measurement of the rate of phototaxis and chlorophyll a content over extended times for up to 12 days. Methodological approaches for analysing time series are discussed, including the recently developed 'GaMoSEC' procedure, providing published references to the detection and understanding of the complex behaviour of natural systems. Finally, we conclude by summarising the general significance of ultradian rhythms to the vital aspects of the biological timekeeping of coherent functionality of living systems from single-cell organisms to humans from their healthy to declining states.
