Proline reduction by<i>Clostridium sporogenes</i>is coupled to vectorial proton ejection

Lovitt, Robert W.; Kell, Douglas B.; Morris, Jonathan

doi:10.1111/j.1574-6968.1986.tb01708.x

Cited by 35 publications

(9 citation statements)

References 22 publications

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“…First proline racemase (PrdF) converts L -proline into D -proline (Wu and Hurdle, 2014). Already in the eighties it was shown that proline reduction is coupled to proton motive force generation (Lovitt et al, 1986). However, the enzyme complex does not reduce ferredoxin and was proposed to directly interact with the membrane-localized, proton/sodium pumping Rnf complex (Jackson et al, 2006).…”

Section: Resultsmentioning

confidence: 99%

Iron Regulation in Clostridioides difficile

et al. 2018

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The response to iron limitation of several bacteria is regulated by the ferric uptake regulator (Fur). The Fur-regulated transcriptional, translational and metabolic networks of the Gram-positive, pathogen Clostridioides difficile were investigated by a combined RNA sequencing, proteomic, metabolomic and electron microscopy approach. At high iron conditions (15 μM) the C. difficile fur mutant displayed a growth deficiency compared to wild type C. difficile cells. Several iron and siderophore transporter genes were induced by Fur during low iron (0.2 μM) conditions. The major adaptation to low iron conditions was observed for the central energy metabolism. Most ferredoxin-dependent amino acid fermentations were significantly down regulated (had, etf, acd, grd, trx, bdc, hbd). The substrates of these pathways phenylalanine, leucine, glycine and some intermediates (phenylpyruvate, 2-oxo-isocaproate, 3-hydroxy-butyryl-CoA, crotonyl-CoA) accumulated, while end products like isocaproate and butyrate were found reduced. Flavodoxin (fldX) formation and riboflavin biosynthesis (rib) were enhanced, most likely to replace the missing ferredoxins. Proline reductase (prd), the corresponding ion pumping RNF complex (rnf) and the reaction product 5-aminovalerate were significantly enhanced. An ATP forming ATPase (atpCDGAHFEB) of the F0F1-type was induced while the formation of a ATP-consuming, proton-pumping V-type ATPase (atpDBAFCEKI) was decreased. The [Fe-S] enzyme-dependent pyruvate formate lyase (pfl), formate dehydrogenase (fdh) and hydrogenase (hyd) branch of glucose utilization and glycogen biosynthesis (glg) were significantly reduced, leading to an accumulation of glucose and pyruvate. The formation of [Fe-S] enzyme carbon monoxide dehydrogenase (coo) was inhibited. The fur mutant showed an increased sensitivity to vancomycin and polymyxin B. An intensive remodeling of the cell wall was observed, Polyamine biosynthesis (spe) was induced leading to an accumulation of spermine, spermidine, and putrescine. The fur mutant lost most of its flagella and motility. Finally, the CRISPR/Cas and a prophage encoding operon were downregulated. Fur binding sites were found upstream of around 20 of the regulated genes. Overall, adaptation to low iron conditions in C. difficile focused on an increase of iron import, a significant replacement of iron requiring metabolic pathways and the restructuring of the cell surface for protection during the complex adaptation phase and was only partly directly regulated by Fur.

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Section: Resultsmentioning

confidence: 99%

Iron Regulation in Clostridioides difficile

et al. 2018

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“…In C. sporogenes , the reduction of D-proline was described to be coupled to transmembrane proton ejection. However, it has not been demonstrated whether this proton gradient is coupled to ATP synthesis [50]. This is possibly due to a very low proton ejection.…”

Section: Resultsmentioning

confidence: 99%

Clostridium sticklandii, a specialist in amino acid degradation:revisiting its metabolism through its genome sequence

et al. 2010

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BackgroundClostridium sticklandii belongs to a cluster of non-pathogenic proteolytic clostridia which utilize amino acids as carbon and energy sources. Isolated by T.C. Stadtman in 1954, it has been generally regarded as a "gold mine" for novel biochemical reactions and is used as a model organism for studying metabolic aspects such as the Stickland reaction, coenzyme-B12- and selenium-dependent reactions of amino acids. With the goal of revisiting its carbon, nitrogen, and energy metabolism, and comparing studies with other clostridia, its genome has been sequenced and analyzed.ResultsC. sticklandii is one of the best biochemically studied proteolytic clostridial species. Useful additional information has been obtained from the sequencing and annotation of its genome, which is presented in this paper. Besides, experimental procedures reveal that C. sticklandii degrades amino acids in a preferential and sequential way. The organism prefers threonine, arginine, serine, cysteine, proline, and glycine, whereas glutamate, aspartate and alanine are excreted. Energy conservation is primarily obtained by substrate-level phosphorylation in fermentative pathways. The reactions catalyzed by different ferredoxin oxidoreductases and the exergonic NADH-dependent reduction of crotonyl-CoA point to a possible chemiosmotic energy conservation via the Rnf complex. C. sticklandii possesses both the F-type and V-type ATPases. The discovery of an as yet unrecognized selenoprotein in the D-proline reductase operon suggests a more detailed mechanism for NADH-dependent D-proline reduction. A rather unusual metabolic feature is the presence of genes for all the enzymes involved in two different CO2-fixation pathways: C. sticklandii harbours both the glycine synthase/glycine reductase and the Wood-Ljungdahl pathways. This unusual pathway combination has retrospectively been observed in only four other sequenced microorganisms.ConclusionsAnalysis of the C. sticklandii genome and additional experimental procedures have improved our understanding of anaerobic amino acid degradation. Several specific metabolic features have been detected, some of which are very unusual for anaerobic fermenting bacteria. Comparative genomics has provided the opportunity to study the lifestyle of pathogenic and non-pathogenic clostridial species as well as to elucidate the difference in metabolic features between clostridia and other anaerobes.

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“…Glycine reduction results in production of acetyl phosphate, and thus ATP synthesis via substrate-level phosphorylation [40, 41]. Reduction of proline has been tied to membrane gradients [42]. The enzymes that catalyze these reactions in C. difficile are glycine reductase and D-proline reductase respectively.…”

Section: Resultsmentioning

confidence: 99%

Auranofin disrupts selenium metabolism in Clostridium difficile by forming a stable Au–Se adduct

et al. 2009

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Clostridium difficile is a nosocomial pathogen whose incidence and importance are on the rise. Previous work in our laboratory characterized the central role of selenoenzyme dependent Stickland reactions in C. difficile metabolism. In this work we have identified, using mass spectrometry, a stable complex formed upon reaction of auranofin (a gold containing drug) with selenide in vitro. X-ray absorption spectroscopy supports the structure that we proposed based on mass spectrometric data. Auranofin potently inhibits the growth of C. difficile but does not similarly affect other clostridia that do not utilize selenoproteins to obtain energy. Moreover, auranofin inhibits the incorporation of radioisotope selenium (75Se) in selenoproteins in both E. coli, the prokaryotic model for selenoprotein synthesis, and C. difficile without impacting total protein synthesis. Auranofin blocks the uptake of selenium and results in the accumulation of the auranofin-selenide adduct in the culture medium. Addition of selenium in the form of selenite or L-selenocysteine to the growth media significantly reduces the inhibitory action of auranofin on the growth of C. difficile. Based on these results, we propose that formation of this complex and the subsequent deficiency in available selenium for selenoprotein synthesis is the mechanism by which auranofin inhibits C. difficile growth. This study demonstrates that targeting selenium metabolism provides a new avenue for antimicrobial development against C. difficile and other selenium-dependent pathogens.

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Proline reduction byClostridium sporogenesis coupled to vectorial proton ejection

Cited by 35 publications

References 22 publications

Iron Regulation in Clostridioides difficile

Iron Regulation in Clostridioides difficile

Clostridium sticklandii, a specialist in amino acid degradation:revisiting its metabolism through its genome sequence

Auranofin disrupts selenium metabolism in Clostridium difficile by forming a stable Au–Se adduct

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