Acetate Metabolism in Archaea: Characterization of an Acetate Transporter and of Enzymes Involved in Acetate Activation and Gluconeogenesis in Haloferax volcanii

Kuprat, Tom; Johnsen, Ulrike; Ortjohann, Marius; Schönheit, Peter

doi:10.3389/fmicb.2020.604926

Cited by 7 publications

(11 citation statements)

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“…A potential candidate might be PEP synthetase for which in the thermophilic archaeon Thermococcus kodakarensis a glycolytic function of PEP conversion to pyruvate has been proposed (36). However, in H. volcanii, a significant contribution of PEP synthetase in glucose degradation seems unlikely since a PEP synthetase deletion mutant was previously shown to grow on glucose as the wild type (12).…”

Section: Resultsmentioning

confidence: 99%

“…In halophilic archaea, sugar metabolism has been studied in particular in Haloferax and Haloarcula species (5)(6)(7)(8)(9)(10). Haloferax volcanii, which represents a model organism of haloarchaea (11), exhibits a high metabolic versatility in utilizing various sugars, hexoses, pentoses and deoxysugars, and other carbon compounds, e.g., acetate, as growth substrates (12). Glucose degradation in H. volcanii has been shown to proceed via the semiphosphorylative ED (spED) pathway, and the enzymes of this pathway, involved in the conversion of glucose to 3-phosphoglycerate (3PG), have been characterized (7,13,86).…”

Section: Importancementioning

confidence: 99%

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Glucose Metabolism and Acetate Switch in Archaea: the Enzymes in Haloferax volcanii

et al. 2021

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The halophilic archaeon Haloferax volcanii has been proposed to degrade glucose via the semiphosphorylative Entner-Doudoroff (spED) pathway. Following our previous studies on key enzymes of this pathway, we now focus on the characterization of enzymes involved in 3-phosphoglycerate conversion to pyruvate, in anaplerosis and in acetyl-CoA formation from pyruvate. These enzymes include phosphoglycerate mutase, enolase, pyruvate kinase, phosphoenolpyruvate carboxylase and pyruvate: ferredoxin oxidoreductase. The essential function of these enzymes were shown by transcript analyses and growth experiments with respective deletion mutants. Further, it is shown that H. volcanii - during aerobic growth on glucose - excreted significant amounts of acetate, which was consumed in the stationary phase (acetate switch). The enzyme catalyzing the conversion of acetyl-CoA to acetate as part of the acetate overflow mechanism, an ADP-forming acetyl-CoA synthetase (ACD), was characterized. The functional involvement of ACD in acetate formation and of AMP-forming acetyl-CoA synthetases (ACSs) in activation of excreted acetate was proven by using respective deletion mutants. Together, the data provide a comprehensive analysis of enzymes of the spED pathway, of anaplerosis, and report first genetic evidence of the functional involvement of enzymes of the acetate switch in archaea. Importance: In this work we provide a comprehensive analysis of glucose degradation via the semiphosphorylative Entner-Doudoroff pathway in the haloarchaeal model organism Haloferax volcanii. The study includes transcriptional analyses, growth experiments with deletion mutants and characterization of all enzymes involved in the conversion of 3-phosphoglycerate to acetyl-CoA and in anaplerosis. Phylogenetic analyses of several enzymes indicate various lateral gene transfer events from bacteria to haloarchaea. Further, we analyzed the key players involved in the acetate switch, i.e in the formation (overflow) and subsequent consumption of acetate during aerobic growth on glucose. Together, the data provide novel aspects on glucose degradation, anaplerosis and acetate switch in H. volcanii and thus expand our understanding of the unusual sugar metabolism in archaea.

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Section: Resultsmentioning

confidence: 99%

Section: Importancementioning

confidence: 99%

Glucose Metabolism and Acetate Switch in Archaea: the Enzymes in Haloferax volcanii

et al. 2021

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“…The activities of isocitrate lyase and malate synthase in cell extracts of Haloferax volcanii , a halophilic archaeon capable of growing in minimal medium with acetate as the only carbon source, revealed no activity when lactate was the carbon source. On the contrary, when acetate was the sole carbon source, the activity of both the enzymes was high ( Serrano et al, 1998 ; Kuprat et al, 2020 ). Cryptococcus neoformans pulmonary infection studies revealed the expression of genes encoding glyoxylate pathway functions (e.g., isocitrate lyase) – those required for growth on acetate – for full virulence.…”

Section: Discussionmentioning

confidence: 99%

Metabolomic Profile of the Fungus Cryomyces antarcticus Under Simulated Martian and Space Conditions as Support for Life-Detection Missions on Mars

et al. 2022

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The identification of traces of life beyond Earth (e.g., Mars, icy moons) is a challenging task because terrestrial chemical-based molecules may be destroyed by the harsh conditions experienced on extraterrestrial planetary surfaces. For this reason, studying the effects on biomolecules of extremophilic microorganisms through astrobiological ground-based space simulation experiments is significant to support the interpretation of the data that will be gained and collected during the ongoing and future space exploration missions. Here, the stability of the biomolecules of the cryptoendolithic black fungus Cryomyces antarcticus, grown on two Martian regolith analogues and on Antarctic sandstone, were analysed through a metabolomic approach, after its exposure to Science Verification Tests (SVTs) performed in the frame of the European Space Agency (ESA) Biology and Mars Experiment (BIOMEX) project. These tests are building a set of ground-based experiments performed before the space exposure aboard the International Space Station (ISS). The analysis aimed to investigate the effects of different mineral mixtures on fungal colonies and the stability of the biomolecules synthetised by the fungus under simulated Martian and space conditions. The identification of a specific group of molecules showing good stability after the treatments allow the creation of a molecular database that should support the analysis of future data sets that will be collected in the ongoing and next space exploration missions.

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“…Its genome has been sequenced and carefully annotated [ 1 , 10 , 11 ]. A plethora of biological aspects have been successfully tackled in this species, with examples including DNA replication [ 4 ]; cell division and cell shape [ 12 , 13 , 14 , 15 , 16 ]; metabolism [ 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 , 25 ]; protein secretion [ 26 , 27 , 28 , 29 ]; motility and biofilms [ 30 , 31 , 32 , 33 , 34 , 35 ]; mating [ 36 ]; signaling [ 37 ]; virus defense [ 38 ]; proteolysis [ 39 , 40 , 41 , 42 , 43 , 44 ]; posttranslational modification (N-glycosylation; SAMPylation) [ 45 , 46 , 47 , 48 , 49 , 50 ]; gene regulation [ 21 , 25 , 51 , 52 , 53 , 54 , 55 ]; microproteins [ 56 , 57 , 58 ] and small noncoding RNAs (sRNAs) […”

Section: Introductionmentioning

confidence: 99%

Open Issues for Protein Function Assignment in Haloferax volcanii and Other Halophilic Archaea

Pfeiffer

Dyall‐Smith

2021

Genes

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Background: Annotation ambiguities and annotation errors are a general challenge in genomics. While a reliable protein function assignment can be obtained by experimental characterization, this is expensive and time-consuming, and the number of such Gold Standard Proteins (GSP) with experimental support remains very low compared to proteins annotated by sequence homology, usually through automated pipelines. Even a GSP may give a misleading assignment when used as a reference: the homolog may be close enough to support isofunctionality, but the substrate of the GSP is absent from the species being annotated. In such cases, the enzymes cannot be isofunctional. Here, we examined a variety of such issues in halophilic archaea (class Halobacteria), with a strong focus on the model haloarchaeon Haloferax volcanii. Results: Annotated proteins of Hfx. volcanii were identified for which public databases tend to assign a function that is probably incorrect. In some cases, an alternative, probably correct, function can be predicted or inferred from the available evidence, but this has not been adopted by public databases because experimental validation is lacking. In other cases, a probably invalid specific function is predicted by homology, and while there is evidence that this assigned function is unlikely, the true function remains elusive. We listed 50 of those cases, each with detailed background information, so that a conclusion about the most likely biological function can be drawn. For reasons of brevity and comprehension, only the key aspects are listed in the main text, with detailed information being provided in a corresponding section of the Supplementary Materials. Conclusions: Compiling, describing and summarizing these open annotation issues and functional predictions will benefit the scientific community in the general effort to improve the evaluation of protein function assignments and more thoroughly detail them. By highlighting the gaps and likely annotation errors currently in the databases, we hope this study will provide a framework for experimentalists to systematically confirm (or disprove) our function predictions or to uncover yet more unexpected functions.

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Acetate Metabolism in Archaea: Characterization of an Acetate Transporter and of Enzymes Involved in Acetate Activation and Gluconeogenesis in Haloferax volcanii

Cited by 7 publications

References 65 publications

Glucose Metabolism and Acetate Switch in Archaea: the Enzymes in Haloferax volcanii

Glucose Metabolism and Acetate Switch in Archaea: the Enzymes in Haloferax volcanii

Metabolomic Profile of the Fungus Cryomyces antarcticus Under Simulated Martian and Space Conditions as Support for Life-Detection Missions on Mars

Open Issues for Protein Function Assignment in Haloferax volcanii and Other Halophilic Archaea

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