Metals in Microbiology
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Metals catalyse almost a half of all microbial reactions and yet can poison microorganisms. Metal handling systems - that maintain metal homeostasis - are thus vital to sustain microbial life. For microbial pathogens, the challenge of metal homeostasis is exacerbated by host immune defences that restrict metal access and that exploit the microbicidal activity of metals. There are opportunities to produce new antimicrobials that subvert microbial metal-handling systems or that use metals directly or combined with other compounds. The prevalence of metalloenzymes also means that engineering the metal-supply in microorganisms is highly relevant to industrial biotechnological processes, with metalloproteins contributing to bioenergy production, bioremediation, biomedicine, synthesis of high value industrial feedstocks and more.
Guest-edited by Dr Jennifer Cavet (University of Manchester) and Dr Karrera Djoko (Durham University), this collection of keynote research articles will highlight research on metal-microbe interactions, bringing together advances in our understanding of how microbes handle metals, the utilization of metals in proteins and the importance of metal handling systems in host-pathogen interactions. It will also include research that exploits these systems in industrial processes, the development of metal-related antimicrobials and in metal bioremediation and biorecovery.
Image credit: Jin Hong
Collection Contents
40 results
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Role of horizontally transferred copper resistance genes in Staphylococcus aureus and Listeria monocytogenes
Bacteria have evolved mechanisms which enable them to control intracellular concentrations of metals. In the case of transition metals, such as copper, iron and zinc, bacteria must ensure enough is available as a cofactor for enzymes whilst at the same time preventing the accumulation of excess concentrations, which can be toxic. Interestingly, metal homeostasis and resistance systems have been found to play important roles in virulence. This review will discuss the copper homeostasis and resistance systems in Staphylococcus aureus and Listeria monocytogenes and the implications that acquisition of additional copper resistance genes may have in these pathogens.
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Application of fungal copper carbonate nanoparticles as environmental catalysts: organic dye degradation and chromate removal
More LessBiomineralization is a ubiquitous process in organisms to produce biominerals, and a wide range of metallic nanoscale minerals can be produced as a consequence of the interactions of micro-organisms with metals and minerals. Copper-bearing nanoparticles produced by biomineralization mechanisms have a variety of applications due to their remarkable catalytic efficiency, antibacterial properties and low production cost. In this study, we demonstrate the biotechnological potential of copper carbonate nanoparticles (CuNPs) synthesized using a carbonate-enriched biomass-free ureolytic fungal spent culture supernatant. The efficiency of the CuNPs in pollutant remediation was investigated using a dye (methyl red) and a toxic metal oxyanion, chromate Cr(VI). The biogenic CuNPs exhibited excellent catalytic properties in a Fenton-like reaction to degrade methyl red, and efficiently removed Cr(VI) from solution due to both adsorption and reduction of Cr(VI). X-ray photoelectron spectroscopy (XPS) identified the oxidation of reducing Cu species of the CuNPs during the reaction with Cr(VI). This work shows that urease-positive fungi can play an important role not only in the biorecovery of metals through the production of insoluble nanoscale carbonates, but also provides novel and simple strategies for the preparation of sustainable nanomineral products with catalytic properties applicable to the bioremediation of organic and metallic pollutants, solely and in mixtures.
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Orchestrating a heist: uptake and storage of metals by apicomplexan parasites
More LessThe acquisition and storage of metals has been a preoccupation of life for millennia. Transition metals, in particular iron, copper and zinc, have vital roles within cells. However, metals also make dangerous cargos; inappropriate uptake or storage of transition metals leads to cell death. This paradox has led to cells developing elegant and frequently redundant mechanisms for fine-tuning local metal concentrations. In the context of infection, pathogens must overcome further hurdles, as hosts act to weaponize metal availability to prevent pathogen colonization and spread. Here, we detail the methods used by the Apicomplexa, a large family of eukaryotic parasites, to obtain and store essential metals.
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At the metal–metabolite interface in Aspergillus fumigatus: towards untangling the intersecting roles of zinc and gliotoxin
Cryptic links between apparently unrelated metabolic systems represent potential new drug targets in fungi. Evidence of such a link between zinc and gliotoxin (GT) biosynthesis in Aspergillus fumigatus is emerging. Expression of some genes of the GT biosynthetic gene cluster gli is influenced by the zinc-dependent transcription activator ZafA, zinc may relieve GT-mediated fungal growth inhibition and, surprisingly, GT biosynthesis is influenced by zinc availability. In A. fumigatus, dithiol gliotoxin (DTG), which has zinc-chelating properties, is converted to either GT or bis-dethiobis(methylthio)gliotoxin (BmGT) by oxidoreductase GliT and methyltransferase GtmA, respectively. A double deletion mutant lacking both GliT and GtmA was previously observed to be hypersensitive to exogenous GT exposure. Here we show that compared to wild-type exposure, exogenous GT and the zinc chelator N,N,N′,N′-tetrakis(2-pyridinylmethyl)−1,2-ethanediamine (TPEN) inhibit A. fumigatus ΔgliTΔgtmA growth, specifically under zinc-limiting conditions, which can be reversed by zinc addition. While GT biosynthesis is evident in zinc-depleted medium, addition of zinc (1 µM) suppressed GT and activated BmGT production. In addition, secretion of the unferrated siderophore, triacetylfusarinine C (TAFC), was evident by A. fumigatus wild-type (at >5 µM zinc) and ΔgtmA (at >1 µM zinc) in a low-iron medium. TAFC secretion suggests that differential zinc-sensing between both strains may influence fungal Fe3+ requirement. Label-free quantitative proteomic analysis of both strains under equivalent differential zinc conditions revealed protein abundance alterations in accordance with altered metabolomic observations, in addition to increased GliT abundance in ΔgtmA at 5 µM zinc, compared to wild-type, supporting a zinc-sensing deficiency in the mutant strain. The relative abundance of a range of oxidoreductase- and secondary metabolism-related enzymes was also evident in a zinc- and strain-dependent manner. Overall, we elaborate new linkages between zinc availability, natural product biosynthesis and oxidative stress homeostasis in A. fumigatus.
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Key carboxylate residues for iron transit through the prokaryotic ferritin SynFtn
More LessFerritins are proteins forming 24meric rhombic dodecahedral cages that play a key role in iron storage and detoxification in all cell types. Their function requires the transport of Fe2+ from the exterior of the protein to buried di-iron catalytic sites, known as ferroxidase centres, where Fe2+ is oxidized to form Fe3+-oxo precursors of the ferritin mineral core. The route of iron transit through animal ferritins is well understood: the Fe2+ substrate enters the protein via channels at the threefold axes and conserved carboxylates on the inner surface of the protein cage have been shown to contribute to transient binding sites that guide Fe2+ to the ferroxidase centres. The routes of iron transit through prokaryotic ferritins are less well studied but for some, at least, there is evidence that channels at the twofold axes are the major route for Fe2+ uptake. SynFtn, isolated from the cyanobacterium Synechococcus CC9311, is an atypical prokaryotic ferritin that was recently shown to take up Fe2+ via its threefold channels. However, the transfer site carboxylate residues conserved in animal ferritins are absent, meaning that the route taken from the site of iron entry into SynFtn to the catalytic centre is yet to be defined. Here, we report the use of a combination of site-directed mutagenesis, absorbance-monitored activity assays and protein crystallography to probe the effect of substitution of two residues potentially involved in this pathway. Both Glu141 and Asp65 play a role in guiding the Fe2+ substrate to the ferroxidase centre. In the absence of Asp65, routes for Fe2+ to, and Fe3+ exit from, the ferroxidase centre are affected resulting in inefficient formation of the mineral core. These observations further define the iron transit route in what may be the first characterized example of a new class of ferritins peculiar to cyanobacteria.
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Iron influences the expression of colonization factor CS6 of enterotoxigenic Escherichia coli
More LessEnterotoxigenic Escherichia coli (ETEC) is a major pathogen of acute watery diarrhoea. The pathogenicity of ETEC is linked to adherence to the small intestine by colonization factors (CFs) and secretion of heat-labile enterotoxin (LT) and/or heat-stable enterotoxin (ST). CS6 is one of the most common CFs in our region and worldwide. Iron availability functions as an environmental cue for enteropathogenic bacteria, signalling arrival within the human host. Therefore, iron could modify the expression of CS6 in the intestine. The objective of this study was to determine the effect of iron availability on CS6 expression in ETEC. This would help in understanding the importance of iron during ETEC pathogenesis. ETEC strain harbouring CS6 was cultured under increasing concentrations of iron salt to assess the effect on CS6 RNA expression by quantitative RT-PCR, protein expression by ELISA, promoter activity by β-galactosidase activity, and epithelial adhesion on HT-29 cells. RNA expression of CS6 was maximum in presence of 0.2 mM iron (II) salt. The expression increased by 50-fold, which also reduced under iron-chelation conditions and an increased iron concentration of 0.4 mM or more. The surface expression of CS6 also increased by 60-fold in presence of 0.2 mM iron. The upregulation of CS6 promoter activity by 25-fold under this experimental condition was in accordance with the induction of CS6 RNA and protein. This increased CS6 expression was independent of ETEC strains. Bacterial adhesion to HT-29 epithelial cells was also enhanced by five-fold in the presence of 0.2 mM iron salt. These findings suggest that CS6 expression is dependent on iron concentration. However, with further increases in iron concentration beyond 0.2 mM CS6 expression is decreased, suggesting that there might be a strong regulatory mechanism for CS6 expression under different iron concentrations.
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Idification and structural characterisation of a catecholate-type siderophore produced by Stenotrophomonas maltophilia K279a
More LessSiderophores are produced by several bacteria that utilise iron in various environments. Elucidating the structure of a specific siderophore may have valuable applications in drug development. Stenotrophomonas maltophilia , a Gram-negative bacterium that inhabits a wide range of environments and can cause pneumonia, produces siderophores. However, the structure was unknown, and therefore, in this study, we aimed to elucidate it. We purified siderophores from cultures of S. maltophilia K279a using preparative reversed-phase HPLC. The structure was analysed through LC-MS and 1H and 13C NMR. The results demonstrated that S. maltophilia K279a produces 2,3-dihydroxybenzoylserine (DHBS), a monomer unit of enterobactin. We suggested the uptake of Iron(III) by the DHBS complex. DHBS production by S. maltophilia K279a could be attributed to an incomplete enterobactin pathway. Drugs targeting DHBS synthesis could prevent S. maltophilia infection.
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Interplay between central carbon metabolism and metal homeostasis in mycobacteria and other human pathogens
More LessBacterial nutrition is a fundamental aspect of pathogenesis. While the host environment is in principle nutrient-rich, hosts have evolved strategies to interfere with nutrient acquisition by pathogens. In turn, pathogens have developed mechanisms to circumvent these restrictions. Changing the availability of bioavailable metal ions is a common strategy used by hosts to limit bacterial replication. Macrophages and neutrophils withhold iron, manganese, and zinc ions to starve bacteria. Alternatively, they can release manganese, zinc, and copper ions to intoxicate microorganisms. Metals are essential micronutrients and participate in catalysis, macromolecular structure, and signalling. This review summarises our current understanding of how central carbon metabolism in pathogens adapts to local fluctuations in free metal ion concentrations. We focus on the transcriptomics and proteomics data produced in studies of the iron-sparing response in Mycobacterium tuberculosis , the etiological agent of tuberculosis, and consequently generate a hypothetical model linking trehalose accumulation, succinate secretion and substrate-level phosphorylation in iron-starved M. tuberculosis . This review also aims to highlight a large gap in our knowledge of pathogen physiology: the interplay between metal homeostasis and central carbon metabolism, two cellular processes which are usually studied separately. Integrating metabolism and metal biology would allow the discovery of new weaknesses in bacterial physiology, leading to the development of novel and improved antibacterial therapies.
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Investigating zinc toxicity responses in marine Prochlorococcus and Synechococcus
More LessMarine plastic pollution is a growing concern worldwide and has the potential to impact marine life via leaching of chemicals, with zinc (Zn), a common plastic additive, observed at particularly high levels in plastic leachates in previous studies. At this time, however, little is known regarding how elevated Zn affects key groups of marine primary producers. Marine cyanobacterial genera Prochlorococcus and Synechococcus are considered to be some of the most abundant oxygenic phototrophs on earth, and together contribute significantly to oceanic primary productivity. Here we set out to investigate how two Prochlorococcus (MIT9312 and NATL2A) and two Synechococcus (CC9311 and WH8102) strains, representative of diverse ecological niches, respond to exposure to high Zn concentrations. The two genera showed differences in the timing and degree of growth and physiological responses to elevated Zn levels, with Prochlorococcus strains showing declines in their growth rate and photophysiology following exposure to 27 µg l−1 Zn, while Synechococcus CC9311 and WH8102 growth rates declined significantly on exposure to 52 and 152 µg l−1 Zn, respectively. Differences were also observed in each strain’s capacity to maintain cell wall integrity on exposure to different levels of Zn. Our results indicate that excess Zn has the potential to pose a challenge to some marine picocyanobacteria and highlights the need to better understand how different marine Prochlorococcus and Synechococcus strains may respond to increasing concentrations of Zn in some marine regions.
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Haem toxicity provides a competitive advantage to the clinically relevant Staphylococcus aureus small colony variant phenotype
Microorganisms encounter toxicities inside the host. Many pathogens exist as subpopulations to maximize survivability. Subpopulations of Staphylococcus aureus include antibiotic-tolerant small colony variants (SCVs). These mutants often emerge following antibiotic treatment but can be present in infections prior to antibiotic exposure. We hypothesize that haem toxicity in the host selects for respiration-deficient S. aureus SCVs in the absence of antibiotics. We demonstrate that some but not all respiration-deficient SCV phenotypes are more protective than the haem detoxification system against transient haem exposure, indicating that haem toxicity in the host may contribute to the dominance of menaquinone-deficient and haem-deficient SCVs prior to antibiotic treatment.
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A novel hemA mutation is responsible for a small-colony-variant phenotype in Escherichia coli
More LessWe identified a small colony variant (SCV) of an amoxicillin/clavulanic acid-resistant derivative of a clinical isolate of Escherichia coli from Malawi, which was selected for in vitro in a subinhibitory concentration of gentamicin. The SCV was auxotrophic for hemin and had impaired biofilm formation compared to the ancestral isolates. A single novel nucleotide polymorphism (SNP) in hemA, which encodes a glutamyl-tRNA reductase that catalyses the initial step of porphyrin biosynthesis leading to the production of haem, was responsible for the SCV phenotype. We showed the SNP in hemA resulted in a significant fitness cost to the isolate, which persisted even in the presence of hemin. However, the phenotype quickly reverted during sequential sub-culturing in liquid growth media. As hemA is not found in mammalian cells, and disruption of the gene results in a significant fitness cost, it represents a potential target for novel drug development specifically for the treatment of catheter-associated urinary tract infections caused by E. coli .
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Ribosome hibernation: a new molecular framework for targeting nonreplicating persisters of mycobacteria
Treatment of tuberculosis requires a multi-drug regimen administered for at least 6 months. The long-term chemotherapy is attributed in part to a minor subpopulation of nonreplicating Mycobacterium tuberculosis cells that exhibit phenotypic tolerance to antibiotics. The origins of these cells in infected hosts remain unclear. Here we discuss some recent evidence supporting the hypothesis that hibernation of ribosomes in M. tuberculosis, induced by zinc starvation, could be one of the primary mechanisms driving the development of nonreplicating persisters in hosts. We further analyse inconsistencies in previously reported studies to clarify the molecular principles underlying mycobacterial ribosome hibernation.
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Molecular basis of bile-salt- and iron-induced enterohaemorrhagic E. coli resistance to cationic antimicrobial peptides
More LessColonization of the gastrointestinal tract by enterohaemorrhagic Escherichia coli (EHEC) is critically dependent on its ability to sense and respond to various microenvironments within the host. EHEC exposure to physiologically relevant levels of bile salts upregulates the two-component system, pmrAB, and the arnBCADTEF operon, resulting in lipopolysaccharide modification and increased resistance to the cationic antimicrobial peptide, polymyxin B (PMB). A similar pmrAB- and arn-dependent PMB resistance has been observed in Salmonella enterica in the presence of ferric iron. Limiting magnesium levels and mild acid can also induce Salmonella resistance to PMB through another two-component system, PhoPQ and the connector protein, PmrD. This study aims to evaluate the relative contributions of a bile-salt mix (BSM), iron, limiting magnesium as well as the roles of pmrAB, phoPQ and pmrD to EHEC’s resistance to PMB. Killing assays show that EHEC treatment with the BSM or iron under excess magnesium and neutral pH conditions induces a pmrAB-dependent, phoP-independent PMB resistance. By contrast, exposure to limiting magnesium triggers a pmrB-, phoP- and pmrD-dependent PMB resistance. The iron-induced PMB resistance is independent of phoP and pmrD under limiting magnesium conditions while the bile-salt-induced PMB resistance is independent of pmrD only under non-PhoP-inducing conditions. GFP-pmrD transcriptional reporter studies reveal that the limiting magnesium enhances pmrD expression, which is repressed upon additional exposure to either BSM or iron. Our results also show that exposure to mild acid enhances PMB resistance in a pmrD-independent manner and GFP reporter results confirm minimal expression of pmrD at this pH regardless of the magnesium level. This study provides novel insights into how EHEC differentially employs PmrAB, PhoPQ and PmrD to monitor and respond to bile salts, iron, acidic pH and magnesium typically encountered within the gastrointestinal tract in order to modulate its survival against cationic antimicrobial peptides.
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Siderophore piracy enhances Vibrio cholerae environmental survival and pathogenesis
More LessVibrio cholerae, the aetiological agent of cholera, possesses multiple iron acquisition systems, including those for the transport of siderophores. How these systems benefit V. cholerae in low-iron, polymicrobial communities in environmental settings or during infection remains poorly understood. Here, we demonstrate that in iron-limiting conditions, co-culture of V. cholerae with a number of individual siderophore-producing microbes significantly promoted V. cholerae growth in vitro. We further show that in the host environment with low iron, V. cholerae colonizes better in adult mice in the presence of the siderophore-producing commensal Escherichia coli . Taken together, our results suggest that in aquatic reservoirs or during infection, V. cholerae may overcome environmental and host iron restriction by hijacking siderophores from other microbes.
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nosX is essential for whole-cell N2O reduction in Paracoccus denitrificans but not for assembly of copper centres of nitrous oxide reductase
Nitrous oxide (N2O) is a potent greenhouse gas that is produced naturally as an intermediate during the process of denitrification carried out by some soil bacteria. It is consumed by nitrous oxide reductase (N2OR), the terminal enzyme of the denitrification pathway, which catalyses a reduction reaction to generate dinitrogen. N2OR contains two important copper cofactors (CuA and CuZ centres) that are essential for activity, and in copper-limited environments, N2OR fails to function, contributing to rising levels of atmospheric N2O and a major environmental challenge. Here we report studies of nosX, one of eight genes in the nos cluster of the soil dwelling α-proteobaterium Paraccocus denitrificans. A P. denitrificans ΔnosX deletion mutant failed to reduce N2O under both copper-sufficient and copper-limited conditions, demonstrating that NosX plays an essential role in N2OR activity. N2OR isolated from nosX-deficient cells was found to be unaffected in terms of the assembly of its copper cofactors, and to be active in in vitro assays, indicating that NosX is not required for the maturation of the enzyme; in particular, it plays no part in the assembly of either of the CuA and CuZ centres. Furthermore, quantitative Reverse Transcription PCR (qRT-PCR) studies showed that NosX does not significantly affect the expression of the N2OR-encoding nosZ gene. NosX is a homologue of the FAD-binding protein ApbE from Pseudomonas stutzeri , which functions in the flavinylation of another N2OR accessory protein, NosR. Thus, it is likely that NosX is a system-specific maturation factor of NosR, and so is indirectly involved in maintaining the reaction cycle of N2OR and cellular N2O reduction.
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Activation of a [NiFe]-hydrogenase-4 isoenzyme by maturation proteases
More LessMaturation of [NiFe]-hydrogenases often involves specific proteases responsible for cleavage of the catalytic subunits. Escherichia coli HycI is the protease dedicated to maturation of the Hydrogenase-3 isoenzyme, a component of formate hydrogenlyase-1. In this work, it is demonstrated that a Pectobacterium atrosepticum HycI homologue, HyfK, is required for hydrogenase-4 activity, a component of formate hydrogenlyase-2, in that bacterium. The P. atrosepticum ΔhyfK mutant phenotype could be rescued by either P. atrosepticum hyfK or E. coli hycI on a plasmid. Conversely, an E. coli ΔhycI mutant was complemented by either E. coli hycI or P. atrosepticum hyfK in trans. E. coli is a rare example of a bacterium containing both hydrogenase-3 and hydrogenase-4, however the operon encoding hydrogenase-4 has no maturation protease gene. This work suggests HycI should be sufficient for maturation of both E. coli formate hydrogenlyases, however no formate hydrogenlyase-2 activity was detected in any E. coli strains tested here.
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Genetic regulation, biochemical properties and physiological importance of arginase from Sinorhizobium meliloti
In bacteria, l-arginine is a precursor of various metabolites and can serve as a source of carbon and/or nitrogen. Arginine catabolism by arginase, which hydrolyzes arginine to l-ornithine and urea, is common in nature but has not been studied in symbiotic nitrogen-fixing rhizobia. The genome of the alfalfa microsymbiont Sinorhizobium meliloti 1021 has two genes annotated as arginases, argI1 (smc03091) and argI2 (sma1711). Biochemical assays with purified ArgI1 and ArgI2 (as 6His-Sumo-tagged proteins) showed that only ArgI1 had detectable arginase activity. A 1021 argI1 null mutant lacked arginase activity and grew at a drastically reduced rate with arginine as sole nitrogen source. Wild-type growth and arginase activity were restored in the argI1 mutant genetically complemented with a genomically integrated argI1 gene. In the wild-type, arginase activity and argI1 transcription were induced several fold by exogenous arginine. ArgI1 purified as a 6His-Sumo-tagged protein had its highest in vitro enzymatic activity at pH 7.5 with Ni2+ as cofactor. The enzyme was also active with Mn2+ and Co2+, both of which gave the enzyme the highest activities at a more alkaline pH. The 6His-Sumo-ArgI1 comprised three identical subunits based on the migration of the urea-dissociated protein in a native polyacrylamide gel. A Lrp-like regulator (smc03092) divergently transcribed from argI1 was required for arginase induction by arginine or ornithine. This regulator was designated ArgIR. Electrophoretic mobility shift assays showed that purified ArgIR bound to the argI1 promoter in a region preceding the predicted argI1 transcriptional start. Our results indicate that ArgI1 is the sole arginase in S. meliloti , that it contributes substantially to arginine catabolism in vivo and that argI1 induction by arginine is dependent on ArgIR.
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Effect of four kinds of complexing iron on the process of iron uptake by Anabaena flos-aquae
More LessAbstract
Iron (Fe), which is a necessary micronutrient for algal growth, plays an important role in the physiological metabolism and enzymatic reactions of algae. This study aimed to investigate the absorption process of four kinds of complexing iron absorbed by Anabaena flos-aquae. Results showed that the absorptive capacity of A. flos-aquae to complex iron was inversely proportional to the stability of the complex bond of complex iron. Complex iron with weak binding ability can be quickly adsorbed by A. flos-aquae. The absorptive rate was as follows: ferric humate, ferric oxalate >ammonium ferric citrate >EDTA Fe. For EDTA-Fe with a strong binding ability, a moderate iron concentration (e.g. 0.6 mg l−1) is favourable for iron uptake by A. flos-aquae. Our experiments also revealed that the process of separating iron from complex iron before entering algal cells was probably as follows: iron complexed with organic ligands were firstly adsorbed on the surface of algae cells; afterwards, iron ions were captured by organic matter on the surface of algae cells, accompanied by the rupture of the bond between Fe3+ and ligand; finally, the Fe3+ entered into the cell of algae while the organic ligands returned to the medium.
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SufT is required for growth of Mycobacterium smegmatis under iron limiting conditions
More LessIron-sulphur (FeS) clusters are versatile cofactors required for a range of biological processes within cells. Due to the reactive nature of the constituent molecules, assembly and delivery of these cofactors requires a multi-protein machinery in vivo. In prokaryotes, SufT homologues are proposed to function in the maturation and transfer of FeS clusters to apo-proteins. This study used targeted gene deletion to investigate the role of SufT in the physiology of mycobacteria, using Mycobacterium smegmatis as a model organism. Deletion of the sufT gene in M. smegmatis had no impact on growth under standard culture conditions and did not significantly alter activity of the FeS cluster dependent enzymes succinate dehydrogenase (SDH) and aconitase (ACN). Furthermore, the ΔsufT mutant was no more sensitive than the wild-type strain to the redox cycler 2,3-dimethoxy-1,4-naphthoquinone (DMNQ), or the anti-tuberculosis drugs isoniazid, clofazimine or rifampicin. In contrast, the ΔsufT mutant displayed a growth defect under iron limiting conditions, and an increased requirement for iron during biofilm formation. This data suggests that SufT is an accessory factor in FeS cluster biogenesis in mycobacteria which is required under conditions of iron limitation.
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Novel intermicrobial molecular interaction: Pseudomonas aeruginosa Quinolone Signal (PQS) modulates Aspergillus fumigatus response to iron
Pseudomonas aeruginosa (Pa) and Aspergillus fumigatus (Af), the commonest bacterium and fungus in compromised host airways, compete for iron (Fe). The Pseudomonas quinolone signal (PQS), a Pa quorum sensing molecule, also chelates Fe, and delivers Fe to the Pa cell membrane using Pa siderophores. In models of Af biofilm formation or preformed biofilms, PQS inhibited Af in a low Fe environment. AfΔsidA (mutant unable to produce siderophores) biofilm was more sensitive to PQS inhibition than wild-type (WT), as was planktonic AfΔsidA growth. PQS decreased WT Af growth on agar. All these inhibitory actions were reversed by Fe. The Pa siderophore pyoverdin, or Af siderophore inhibitor celastrol, act cooperatively with PQS in Af inhibition. These findings all indicate PQS inhibition is owing to Fe chelation. Remarkably, in high Fe environments, PQS enhanced Af biofilm at 1/100 to 1/2000 Fe concentration required for Fe alone to enhance. Planktonic Af growth, and on agar, Af conidiation, were also enhanced by PQS+Fe compared to Fe alone. In contrast, neither AfΔsidA biofilm, nor planktonic AfΔsidA, were enhanced by PQS-Fe compared to Fe. When Af siderophore ferricrocin (FC),+PQS, were added to AfΔsidA, Af was then boosted more than by FC alone. Moreover, FC+PQS+Fe boosted AfΔsidA more than Fe, FC, FC+Fe, PQS+FC or PQS+Fe. Thus PQS-Fe maximal stimulation requires Af siderophores. PQS inhibits Af via chelation under low Fe conditions. In a high Fe environment, PQS paradoxically stimulates Af efficiently, and this involves Af siderophores. PQS production by Pa could stimulate Af in cystic fibrosis airways, where Fe homeostasis is altered and Fe levels increase, supporting fungal growth.
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Shotgun proteomic analysis of nanoparticle-synthesizing Desulfovibrio alaskensis in response to platinum and palladium
More LessPlatinum and palladium are much sought-after metals of critical global importance in terms of abundance and availability. At the nano-scale these metals are of even higher value due to their catalytic abilities for industrial applications. Desulfovibrio alaskensis is able to capture ionic forms of both of these metals, reduce them and synthesize elemental nanoparticles. Despite this ability, very little is known about the biological pathways involved in the formation of these nanoparticles. Proteomic analysis of D. alaskensis in response to platinum and palladium has highlighted those proteins involved in both the reductive pathways and the wider stress-response system. A core set of 13 proteins was found in both treatments and consisted of proteins involved in metal transport and reduction. There were also seven proteins that were specific to either platinum or palladium. Overexpression of one of these platinum-specific genes, a NiFe hydrogenase small subunit (Dde_2137), resulted in the formation of larger nanoparticles. This study improves our understanding of the pathways involved in the metal resistance mechanism of Desulfovibrio and is informative regarding how we can tailor the bacterium for nanoparticle production, enhancing its application as a bioremediation tool and as a way to capture contaminant metals from the environment.
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Inactivation of the Agrobacterium tumefaciens ActSR system affects resistance to multiple stresses with increased H2O2 sensitivity due to reduced expression of hemH
More LessThe Agrobacterium tumefaciens ActSR two-component regulatory system is a member of a homologous group of global redox-responsive regulatory systems that adjust the expression of energy-consuming and energy-supplying metabolic pathways in order to maintain cellular redox balance. In this study, the transcriptional organization of the hrpB-actSR locus was determined and the effect of actSR system inactivation on stress resistance was investigated. It was found that hrpB is transcribed as a monocistronic mRNA and actS is transcribed along with actR as a bicistronic mRNA, while actR is also transcribed as a monocistronic message. Each message is initiated from a separate promoter. Inactivation of actR resulted in decreased resistance to membrane stress (sodium dodecyl sulfate), acid stress (pH 5.5), iron starvation (bipyridyl) and iron excess (FeCl3), and antibiotic stress (tetracycline and ciprofloxacin). Resistance to oxidative stress in the form of organic peroxide (cumene hydroperoxide) increased, while resistance to inorganic peroxide (H2O2) decreased. An actR insertion mutant displayed reduced catalase activity, even though transcription of katA and catE remained unchanged. Complementation of the actR inactivation mutant with plasmid-encoded actR or overexpression of hemH, encoding ferrochelatase, restored wild-type catalase activity and H2O2 resistance levels. Gel mobility shift and hemH promoter–lacZ fusion results indicated that ActR is a positive regulator of hemH that binds directly to the hemH promoter region. Thus, inactivation of the A. tumefaciens ActSR system affects resistance to multiple stresses, including reduced resistance to H2O2 resulting from a reduction in catalase activity due to reduced expression of hemH.
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The regulation of iron homeostasis in the fungal human pathogen Candida glabrata
More LessIron is an essential element to most microorganisms, yet an excess of iron is toxic. Hence, living cells have to maintain a tight balance between iron uptake and iron consumption and storage. The control of intracellular iron concentrations is particularly challenging for pathogens because mammalian organisms have evolved sophisticated high-affinity systems to sequester iron from microbes and because iron availability fluctuates among the different host niches. In this review, we present the current understanding of iron homeostasis and its regulation in the fungal pathogen Candida glabrata. This yeast is an emerging pathogen which has become the second leading cause of candidemia, a life-threatening invasive mycosis. C. glabrata is relatively poorly studied compared to the closely related model yeast Saccharomyces cerevisiae or to the pathogenic yeast Candida albicans. Still, several research groups have started to identify the actors of C. glabrata iron homeostasis and its transcriptional and post-transcriptional regulation. These studies have revealed interesting particularities of C. glabrata and have shed new light on the evolution of fungal iron homeostasis.
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Rhizobium tropici CIAT 899 copA gene plays a fundamental role in copper tolerance in both free life and symbiosis with Phaseolus vulgaris
Rhizobium tropici CIAT 899 is a facultative symbiotic diazotroph able to deal with stressful concentrations of metals. Nevertheless the molecular mechanisms involved in metal tolerance have not been elucidated. Copper (Cu2+) is a metal component essential for the heme-copper respiratory oxidases and enzymes that catalyse redox reactions, however, it is highly toxic when intracellular trace concentrations are surpassed. In this study, we report that R. tropici CIAT 899 is more tolerant to Cu2+ than other Rhizobium and Sinorhizobium species. Through Tn5 random mutagenesis we identify a R. tropici mutant strain with a severe reduction in Cu2+ tolerance. The Tn5 insertion disrupted the gene RTCIAT899_CH17575, encoding a putative heavy metal efflux P1B-1-type ATPase designated as copA. Phaseolus vulgaris plants inoculated with the copA::Tn5 mutant in the presence of toxic Cu2+ concentrations showed a drastic reduction in plant and nodule dry weight, as well as nitrogenase activity. Nodules induced by the copA::Tn5 mutant present an increase in H2O2 concentration, lipoperoxidation and accumulate 40-fold more Cu2+ than nodules formed by the wild-type strain. The copA::Tn5 mutant complemented with the copA gene recovered the wild-type symbiotic phenotypes. Therefore, the copA gene is essential for R. tropici CIAT 899 to survive in copper-rich environments in both free life and symbiosis with P. vulgaris plants.
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The aminoglycoside resistance-promoting AmgRS envelope stress-responsive two-component system in Pseudomonas aeruginosa is zinc-activated and protects cells from zinc-promoted membrane damage
More LessExposure of wild-type (WT) Pseudomonas aeruginosa PAO1 to ZnCl2 (Zn) yielded a concentration-dependent increase in depolarization of the cytoplasmic membrane (CM), an indication that this metal is membrane-damaging. Consistent with this, Zn activated the AmgRS envelope stress-responsive two-component system (TCS) that was previously shown to be activated by and to protect P. aeruginosa from the membrane-damaging effects of aminoglycoside (AG) antibiotics. A mutant lacking amgR showed enhanced Zn-promoted CM perturbation and was Zn-sensitive, an indication that the TCS protected cells from the CM-damaging effects of this metal. In agreement with this, a mutant carrying an AmgRS-activating amgS mutation was less susceptible to Zn-promoted CM perturbation and more tolerant of elevated levels of Zn than WT. AG activation of AmgRS is known to drive expression of the AG resistance-promoting mexXY multidrug efflux operon, and while Zn similarly induced mexXY expression this was independent of AmgRS and reliant on a second TCS implicated in mexXY regulation, ParRS. MexXY did not, however, contribute to Zn resistance or protection from Zn-promoted CM damage. Despite its activation of AmgRS and induction of mexXY, Zn had a minimal impact on the AG resistance of WT P. aeruginosa although, given that Zn-tolerant AmgRS-activated amgS mutant strains are AG resistant, there is still the prospect of this metal promoting AG resistance development in this organism.
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Characterization of 4-guanidinobutyrase from Aspergillus niger
More LessArginase is the only fungal ureohydrolase that is well documented in the literature. More recently, a novel route for agmatine catabolism in Aspergillus niger involving another ureohydrolase, 4-guanidinobutyrase (GBase), was reported. We present here a detailed characterization of A. niger GBase – the first fungal (and eukaryotic) enzyme to be studied in detail. A. niger GBase is a homohexamer with a native molecular weight of 336 kDa and an optimal pH of 7.5. Unlike arginase, the Mn2+ enzyme from the same fungus, purified GBase protein is associated with Zn2+ ions. A sensitive fluorescence assay was used to determine its kinetic parameters. GBase acted 25 times more efficiently on 4-guanidinobutyrate (GB) than 3-guanidinopropionic acid (GP). The Km for GB was 2.7±0.4 mM, whereas for GP it was 53.7±0.8 mM. While GB was an efficient nitrogen source, A. niger grew very poorly on GP. Constitutive expression of GBase favoured fungal growth on GP, indicating that GP catabolism is limited by intracellular GBase levels in A. niger. The absence of a specific GPase and the inability of GP to induce GBase expression confine the fungal growth on GP. That GP is a poor substrate for GBase and a very poor nitrogen source for A. niger offers an opportunity to select GBase specificity mutations. Further, it is now possible to compare two distinct ureohydrolases, namely arginase and GBase, from the same organism.
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LcpRVH2 – regulating the expression of latex-clearing proteins in Gordonia polyisoprenivorans VH2
More LessGordonia polyisoprenivorans VH2 harbours two latex clearing proteins, which are responsible for the cleavage of poly(cis-1,4-isoprene) into oligoisoprenes, thereby allowing growth in presence of, e.g. natural rubber. A gene coding for a putative regulator of the TetR-family (lcpR VH2) is located 131 bp upstream of lcp1 VH2. We heterologously expressed lcpR VH2 in Escherichia coli , and purified and characterized the protein with respect to its ability to bind to the operator region of lcp1 VH2. LcpRVH2 forms a dimer in its native state. The size of the dimer was determined to be 52.7 kDa by size exclusion chromatography, whereas the calculated size of a monomer was 24.1 kDa. Electrophoretic mobility shift assays (EMSAs) with the purified protein revealed a shift upon binding to the intergenic region between lcpR VH2 and lcp1 VH2. Within this region, an inverted repeat was identified in silico, probably being the binding site of LcpRVH2. This binding sequence was confirmed by a DNase I footprinting assay. A shift also occurred in EMSAs with this 44 bp sequence only. Interestingly, no regulator was detected upstream of the second lcp (lcp2 VH2). Therefore, we performed EMSA studies with LcpRVH2 and the putative operator region upstream of lcp2 VH2, and discovered by DNase I footprinting another binding sequence upstream of lcp2 VH2. Hence, we concluded that LcpRVH2 binds the operator region of both lcps and, most likely, regulates their expression in G. polyisoprenivorans VH2.
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The B12 receptor BtuB alters the membrane integrity of Caulobacter crescentus
More LessVitamin B12 is one of the most complex biomolecules in nature. Since few organisms can synthesize B12 de novo, most bacteria utilize highly sensitive and specialized transporters to scavenge B12 and its precursors. In Gram-negative bacteria, BtuB is the outer membrane TonB-dependent receptor for B12. In the fresh water bacterium Caulobacter crescentus , btuB is among the most highly expressed genes. In this study, we characterized the function of BtuB in C. crescentus and unveiled a potential new function of this receptor involved in cellular fitness. Under standard minimal or rich growth conditions, we found that supplements of vitamin B12 to cultures of C. crescentus provided no significant advantage in growth rate. Using a B12 methionine auxotroph, we showed that BtuB in C. crescentus is capable of transporting B12 at low pico-molar range. A btuB knockout strain displayed higher sensitivity to detergents and to changes in osmotic pressure compared to the wild-type. Electron micrographs of this knockout strain revealed a morphology defect. The sensitivity observed in the btuB knockout strain was not due to changes in membrane permeability or altered S-layer levels. Our results demonstrate that btuB deletion mutants exhibit increased susceptibility to membrane stressors, suggesting a potential role of this receptor in membrane homeostasis. Because we only tested BtuB’s function under laboratory conditions, we cannot eliminate the possibility that BtuB also plays a key role as a B12 scavenger in C. crescentus when growing in its highly variable and nutrient-limited natural environment.
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The copper-inducible copAB operon in Xanthomonas citri subsp. citri is regulated at transcriptional and translational levels
Upstream open reading frames (ORFs) are frequently found in the 5′-flanking regions of genes and may have a regulatory role in gene expression. A small ORF (named cohL here) was identified upstream from the copAB copper operon in Xanthomonascitri subsp. citri (Xac). We previously demonstrated that copAB expression was induced by copper and that gene inactivation produced a mutant strain that was unable to grow in the presence of copper. Here, we address the role of cohL in copAB expression control. We demonstrate that cohL expression is induced by copper in a copAB-independent manner. Although cohL is transcribed, the CohL protein is either not expressed in vivo or is synthesized at undetectable levels. Inactivation of cohL ( X. citri cohL polar mutant strain) leads to an inability to synthesize cohL and copAB transcripts and consequently the inability to grow in the presence of copper. Bioinformatic tools predicted a stem–loop structure for the cohL–copAB intergenic region and revealed that this region may arrange itself in a secondary structure. Using in vitro gene expression, we found out that the structured 5′-UTR mRNA of copAB is responsible for sequestering the ribosome-binding site that drives the translation of copA. However, copper alone was not able to release the sequence. Based on the results, we speculate that cohL plays a role as a regulatory RNA rather than as a protein-coding gene.
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Effect of limitation of iron and manganese on microalgae growth in fresh water
More LessEutrophication is caused by the rapid growth of microalgae. Iron and manganese are important micronutrients for microalgae growth. However, the effect of the limitation of iron and manganese on microalgae growth in fresh water has not been well understood. In this study, natural mixed algae, Anabaena flosaquae and Scenedesmus quadricanda, were cultivated under different quotas of iron and manganese to reveal the effect of the limitation of iron and manganese on the growth of microalgae in fresh water. The results showed that the growth rate of algae is influenced more by iron than by manganese. However, the effect of manganese cannot be overlooked: when the initial manganese quota was replete, i.e. 0.6–0.8 mg l−1, manganese was able to relieve the effect of iron limitation on microalgae growth in fresh water. We further found that the microalgae showed an uptake preference for iron over manganese. Iron had a competitive effect on manganese uptake, while manganese had less impact on iron uptake by microalgae. The information obtained in the current study is useful for the provision of water quality warnings and for the control of microalgae bloom in fresh water.
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The monofunctional cobalamin biosynthesis enzyme precorrin-3B synthase (CobZRR) is essential for anaerobic photosynthesis in Rhodospirillum rubrum but not for aerobic dark metabolism
More LessThe in vivo physiological role of the gene cobZ, which encodes precorrin-3B synthase, which catalyzes the initial porphyrin ring contraction step of cobalamin biosynthesis via the cob pathway, has been demonstrated here for the first time. Cobalamin is known to be essential for an early step of bacteriochlorophyll biosynthesis in anoxygenic purple bacteria. The cobZ (cobZRR ) gene of the purple bacterium Rhodospirillum rubrum was localized to a 23.5 kb insert of chromosomal DNA contained on the cosmid pSC4. pSC4 complemented several mutants of bacteriochlorophyll and carotenoid biosynthesis, due to the presence of the bchCX and crtCDEF genes at one end of the cosmid insert, flanking cobZRR . A second gene, citB/tcuB, immediately downstream of cobZRR , shows homologies to both a tricarballylate oxidoreductase (tcuB) and a gene (citB) involved in signal transduction during citrate uptake. CobZRR shows extensive homology to the N-terminal domain of the bifunctional CobZ from Rhodobacter capsulatus, and the R. rubrum citB/tcuB gene is homologous to the CobZ C-terminal domain. A mutant, SERGK25, containing a terminatorless kanamycin interposon inserted into cobZRR , could not grow by anaerobic photosynthesis, but grew normally under dark, aerobic and microaerophilic conditions with succinate and fructose as carbon sources. The anaerobic in vivo activity of CobZ indicates that it does not require oxygen as a substrate. The mutant excreted large amounts of protoporphyrin IX-monomethylester, a brown precursor of bacteriochlorophyll biosynthesis. The mutant was complemented either by the cobZRR gene in trans, or when exogenous cobalamin was added to the medium. A deletion mutant of tcuB/citB did not exhibit the cob phenotype. Thus, a role for tcuB/citB in cobalamin biosynthesis could not be confirmed.
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Defining the regulatory mechanism of NikR, a nickel-responsive transcriptional regulator, in Brucella abortus
More LessMetals are essential micronutrients for virtually all forms of life, but metal acquisition is a double-edged sword, because high concentrations of divalent cations can be toxic to the cell. Therefore, the genes involved in metal acquisition, storage and efflux are tightly regulated. The present study characterizes a nickel-responsive transcriptional regulator in the intracellular mammalian pathogen, Brucella abortus. Deletion of bab2_0432 (nikR) in B. abortus led to alterations in the nickel-responsive expression of the genes encoding the putative nickel importer NikABCDE and, moreover, NikR binds directly to a specific DNA sequence within the promoter region of nikA in a metal-dependent manner to control gene expression. While NikR is involved in controlling the expression of nikA, nikR is not required for the infection of macrophages or mice by B. abortus. Overall, this work characterizes the role of NikR in nickel-responsive gene expression, as well as the dispensability of nikR for Brucella virulence.
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Manganese is a Deinococcus radiodurans growth limiting factor in rich culture medium
To understand the effects triggered by Mn2+ on Deinococcus radiodurans, the proteome patterns associated with different growth phases were investigated. In particular, under physiological conditions we tested the growth rate and the biomass yield of D. radiodurans cultured in rich medium supplemented or not with MnCl2. The addition of 2.5–5.0 µM MnCl2 to the medium neither altered the growth rate nor the lag phase, but significantly increased the biomass yield. When higher MnCl2 concentrations were used (10–250 µM), biomass was again found to be positively affected, although we did observe a concentration-dependent lag phase increase. The in vivo concentration of Mn2+ was determined in cells grown in rich medium supplemented or not with 5 µM MnCl2. By atomic absorption spectroscopy, we estimated 0.2 and 0.75 mM Mn2+ concentrations in cells grown in control and enriched medium, respectively. We qualitatively confirmed this observation using a fluorescent turn-on sensor designed to selectively detect Mn2+ in vivo. Finally, we investigated the proteome composition of cells grown for 15 or 19 h in medium to which 5 µM MnCl2 was added, and we compared these proteomes with those of cells grown in the control medium. The presence of 5 µM MnCl2 in the culture medium was found to alter the pI of some proteins, suggesting that manganese affects post-translational modifications. Further, we observed that Mn2+ represses enzymes linked to nucleotide recycling, and triggers overexpression of proteases and enzymes linked to the metabolism of amino acids.
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Helicobacter pylori nickel storage proteins: recognition and modulation of diverse metabolic targets
More LessNickel metabolism and trafficking in Helicobacter pylori is complex, perhaps more so than in any other pathogen. Along with nickel enzymes and their associated nickel-binding maturation machinery, H. pylori contains nickel storage proteins, Hpn and Hpnl. Through a combined crosslinking and enrichment approach, we show that Hpn/Hpnl interact with a wide array of partners; over 100 proteins were captured, including known nickel-enzyme maturation proteins, and other proteins outside known H. pylori nickel-associated proteins. The crosslinker binds to exposed amines, but there was no correlation between lysine content and the pulldown abundance of captured proteins. Phenotypic characterization of mutant strains (Δhpn, Δhpnl, or ΔhpnΔhpnl) was used to explore interactions. Nickel deprivation affected the hydrogenase activity of the ΔhpnΔhpnl strain much more severely than the wild-type (WT), whereas the activities of the single mutants were similar to WT. Leucyl aminopeptidase activity was affected in opposite ways in the mutant strains: Δhpn had a threefold decrease, while Δhpnl had a sevenfold increase, compared to the parent. Similar mutant strain analysis supported Hpn and Hpnl acting synergistically to suppress aliphatic amidase activity in a nickel-dependent manner. Recombinant amidase could bind a variety of divalent metals. Amidase activity was greatest in the mutant strains and was inhibited by exogenous nickel. The addition of pure storage protein to extracts from the mutants only restored the suppression of amidase activity for the mutant strain lacking that protein; both storage proteins are needed for amidase suppression. These results suggest that Hpn and Hpnl play more diverse roles than previously thought.
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Intergenic evolution during host adaptation increases expression of the metallophore pseudopaline in Pseudomonas aeruginosa
Regulating intracellular levels of biological metal ions is essential for all bacterial species, as they are needed for virulence and a range of metabolic processes. Zinc is the second most abundant metal ion in Pseudomonas aeruginosa, but little is known about its regulation. Recent studies have identified a novel operon, zrmABCD (also called cntOLMI), encoding a metallophore system (pseudopaline) involved in zinc acquisition. Expression of this operon has been implicated in human infections and is regulated by the transcriptional regulator Zur (Zn2+ uptake regulator). In this study, we show that the intergenic promoter region in front of zrmABCD is a target for recurrent adaptive mutations during chronic infection of cystic fibrosis (CF) patients. We characterize the inter- and intraclonal sequence polymorphisms found in the promoter region of the metallophore system and find that most alterations increase promoter activity. One of the evolved promoters displays a more than 10-fold increase compared to the ancestral strain due to the combined effect of an altered binding site of Zur and changes to the RpoD-binding motif. This specific evolved promoter responds differently to changes in metal ion concentrations in chelated medium. We have previously shown that P. aeruginosa evolves toward iron acquisition from haemoglobin during long-term CF infections. We hereby provide the second example of adaptive mutations targeting intergenic regions that affect metal ion uptake systems during CF infections, and the first involving zinc uptake. Our results suggest that the scarcity of metal ions (including iron and zinc) is an important evolutionary driver in CF host adaptation.
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Control of zinc homeostasis in Agrobacterium tumefaciens via zur and the zinc uptake genes znuABC and zinT
The Agrobacterium tumefaciens zinc uptake regulator (Zur) was shown to negatively regulate the zinc uptake genes znuABC, encoding a zinc transport system belonging to the ATP-binding cassette (ABC) transporter family, and zinT, which encodes a periplasmic zinc-binding protein. The expression of znuABC and zinT was inducible when cells were grown in medium containing a metal chelator (EDTA), and this induction was shown to be specific for zinc depletion. The expression of znuABC was reduced in response to increased zinc in a dose-dependent manner, and zinT had a less pronounced but similar pattern of zinc-regulated expression. The inactivation of zur led to constitutively high expression of znuABC and zinT. In addition, a zur mutant had an increased total zinc content compared to the WT NTL4 strain, whereas the inactivation of zinT caused a reduction in the total zinc content. The zinT gene is shown to play a dominant role and to be more important than znuA and znuB for A. tumefaciens survival under zinc deprivation. ZinT can function even when ZnuABC is inactivated. However, mutations in zur, znuA, znuB or zinT did not affect the virulence of A. tumefaciens.
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Metals, minerals and microbes: geomicrobiology and bioremediation
More LessMicrobes play key geoactive roles in the biosphere, particularly in the areas of element biotransformations and biogeochemical cycling, metal and mineral transformations, decomposition, bioweathering, and soil and sediment formation. All kinds of microbes, including prokaryotes and eukaryotes and their symbiotic associations with each other and ‘higher organisms’, can contribute actively to geological phenomena, and central to many such geomicrobial processes are transformations of metals and minerals. Microbes have a variety of properties that can effect changes in metal speciation, toxicity and mobility, as well as mineral formation or mineral dissolution or deterioration. Such mechanisms are important components of natural biogeochemical cycles for metals as well as associated elements in biomass, soil, rocks and minerals, e.g. sulfur and phosphorus, and metalloids, actinides and metal radionuclides. Apart from being important in natural biosphere processes, metal and mineral transformations can have beneficial or detrimental consequences in a human context. Bioremediation is the application of biological systems to the clean-up of organic and inorganic pollution, with bacteria and fungi being the most important organisms for reclamation, immobilization or detoxification of metallic and radionuclide pollutants. Some biominerals or metallic elements deposited by microbes have catalytic and other properties in nanoparticle, crystalline or colloidal forms, and these are relevant to the development of novel biomaterials for technological and antimicrobial purposes. On the negative side, metal and mineral transformations by microbes may result in spoilage and destruction of natural and synthetic materials, rock and mineral-based building materials (e.g. concrete), acid mine drainage and associated metal pollution, biocorrosion of metals, alloys and related substances, and adverse effects on radionuclide speciation, mobility and containment, all with immense social and economic consequences. The ubiquity and importance of microbes in biosphere processes make geomicrobiology one of the most important concepts within microbiology, and one requiring an interdisciplinary approach to define environmental and applied significance and underpin exploitation in biotechnology.
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An extracellular zinc metalloprotease gene of Burkholderia cepacia
More LessBurkholderia cepacia produces at least one extracellular zinc metalloprotease that may be involved in virulence. A B. cepacia zinc metalloprotease gene was cloned using a Burkholderia pseudomallei zinc metalloprotease gene as a probe. The predicted amino acid sequences of these B. cepacia and a B. pseudomallei extracellular zinc metalloproteases indicate that they are similar to the thermolysin-like family of metalloproteases (M4 family of metalloendopeptidases) and they are likely to be secreted via the general secretory pathway. zmpA isogenic mutants were constructed in B. cepacia genomovar III strains Pc715j and K56-2 by insertional inactivation of the zmpA genes. The zmpA mutants produced less protease than the parent strains. The B. cepacia strain K56-2 zmpA mutant was significantly less virulent than its parent strain in a chronic respiratory infection model; however, there was no difference between the virulence of B. cepacia strain Pc715j and a Pc715j zmpA mutant. The results indicate that this extracellular zinc metalloprotease may play a greater role in virulence in some strains of B. cepacia.
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