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Theresa Kouril

About Theresa Kouril:

SEEK ID: https://fairdomhub.org/people/261

Location:  Germany

Expertise: functional protein expression, Glycolysis, Microbiology, Molecular Biology, Systems Biology, Carbohydrate Metabolism of Archaea, carbon metabolism, Biochemistry

Tools: Enzyme assays, Molecular Biology, Biochemistry

ORCID: Not specified

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SulfoSys - Biotec

Within the e:Bio - Innovationswettbewerb Systembiologie (Federal Ministry of Education and Research (BMBF)), the SulfoSYSBIOTECH consortium (10 partners), aim to unravel the complexity and regulation of the carbon metabolic network of the thermoacidophilic archaeon Sulfolobus solfataricus (optimal growth at 80°C and pH 3) in order to provide new catalysts ‘extremozymes’ for utilization in White Biotechnology.

Based on the available S. solfataricus genome scale metabolic model (Ulas et al., 2012)
...

Programme: e:Bio

Public web page: http://www.sulfosys.com/

SulfoSys

Silicon cell model for the central carbohydrate metabolism of the archaeon Sulfolobus solfataricus under temperature variation

Programme: SysMO

Public web page: http://sulfosys.com/

University Duisburg-Essen

Country:  Germany

City: Essen

Web page: https://www.uni-due.de/

Amino acid degradation in Sulfolobus solfataricus P2
SulfoSys - Biotec

Integrated systems biology approach including transcriptome, metabolome, proteome analyses and modelling to elucidate amino acid degradation in S. solfataricus P2.

Snapshots: No snapshots

Phosphoglycerate kinase acts as a futile cycle at high temperature
SulfoSys, SulfoSys - Biotec

The gluconeogenic conversion of 3-phosphoglycerate via 1,3-bisphosphoglycerate to glyceraldehyde-3-phosphate was compared at 30 C and at 70 C. At 30 C it was possible to produce 1,3-bisphosphoglycerate from 3-phosphoglycerate with phosphoglycerate kinase, but at 70 C, 1,3- bisphosphoglycerate was dephosphorylated rapidly to 3-phosphoglycerate, effectively turning the phosphoglycerate kinase into a futile cycle. At both temperatures it was possible to convert 3-phosphoglycerate to glyceraldehyde
...

Snapshots: No snapshots

L-fucose degradation in Sulfolobus solfataricus P2
SulfoSys - Biotec

Integrated systems biology approach including transcriptome, metabolome, biochemistry, proteome analyses and modelling to elucidate the catabolic pathway for L-fucose in S. solfataricus P2.

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Comparison of Sulfolobus solfataricus P2 grown on caseinhydrolysate and D-glucose
SulfoSys - Biotec

To investigate amino acid degradation pathways in Sulfolobus solfataricus transcriptome, proteome and metabolome analyses were performed on cells grown on caseinhydrolysate as carbon source. Cells grown with glucose served as reference condition. Metabolic modelling was used to compare the efficiency of different degradation routes.

Person responsible: Jacqueline Wolf

Snapshots: No snapshots

PGK-GAPDH 30C & 70C
SulfoSys, SulfoSys - Biotec

PGK-GAPDH reactions were studied in vitro at 30 and 70 using yeast or Sso enzymes

SED-ML:
https://jjj.bio.vu.nl/models/experiments/kouril2017_fig4b/simulate
https://jjj.bio.vu.nl/models/experiments/kouril2017_fig4c/simulate

Person responsible: Theresa Kouril

Snapshots: No snapshots

PGK-30C
SulfoSys, SulfoSys - Biotec

PGK reaction at 30 C. Yeast PGK was incubated at 30 C, in the presence or absence of the ATP recycling system, and the conversion of 3 PG to BPG was followed.
SED-ML simulations
Fig. 1A: https://jjj.bio.vu.nl/models/experiments/kouril2017_fig1a/simulate

Person responsible: Jacky Snoep

Snapshots: No snapshots

PGK-70C
SulfoSys, SulfoSys - Biotec

PGK reaction at 70C. Sulfolobus solfataricus PGK was incubated at 70C in presence and absence of an ATP recycling system.
Changes in metabolite concentrations was followed via 31P NMR or enzymatic analyses.

SED-ML:
https://jjj.bio.vu.nl/models/experiments/kouril2017_fig3b/simulate
https://jjj.bio.vu.nl/models/experiments/kouril2017_fig3c/simulate
https://jjj.bio.vu.nl/models/experiments/kouril2017_fig3d/simulate

Person responsible: Theresa Kouril

Snapshots: No snapshots

BPG stability
SulfoSys, SulfoSys - Biotec

BPG produced with yeast PGK was incubated at 70 C,upon which BPG rapidly dephosphorylates to 3PG.
SED-ML:
https://jjj.bio.vu.nl/models/experiments/kouril2017_fig2b/simulate

Person responsible: Jacky Snoep

Snapshots: No snapshots

Comparison of S. solfataricus grown on l-fucose and d-glucose
SulfoSys - Biotec
No description specified

Person responsible: Theresa Kouril

Snapshots: No snapshots

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PGK-GAPDH 70
SulfoSys, SulfoSys - Biotec
No description specified

Contributor: Theresa Kouril

Assay type: Experimental Assay Type

Technology type: Technology Type

Snapshots: No snapshots

PGK-GAPDH 30
SulfoSys, SulfoSys - Biotec
No description specified

Contributor: Theresa Kouril

Assay type: Experimental Assay Type

Technology type: Technology Type

Snapshots: No snapshots

BPG degradation
SulfoSys, SulfoSys - Biotec

BPG degradation at 70C

Contributor: Jacky Snoep

Assay type: Metabolite Concentration

Technology type: Technology Type

Snapshots: No snapshots

PGK 70 data
SulfoSys, SulfoSys - Biotec

Changes in metabolite concentrations were either quantified via 31P NMR or enzymatically

Contributor: Theresa Kouril

Assay type: Experimental Assay Type

Technology type: NMR

Snapshots: No snapshots

PGK 30C data
SulfoSys, SulfoSys - Biotec

Experimental data for the yeast PGK incubations at 30C, with and without recycling of ATP.

Contributor: Jacky Snoep

Assay type: Metabolite Concentration

Technology type: Technology Type

Snapshots: No snapshots

non-enzymatic phosphoenolpyruvate (PEP) degradation at high temperature
SulfoSys, SulfoSys - Biotec
No description specified

Contributor: Theresa Kouril

Assay type: Experimental Assay Type

Technology type: Enzymatic Activity Measurements

Snapshots: No snapshots

Cell free extract activity measurements: L-fuc/d-glc
SulfoSys - Biotec
No description specified

Contributor: Jacqueline Wolf

Assay type: Experimental Assay Type

Technology type: Enzymatic Activity Measurements

Snapshots: No snapshots

Proteome analysis: d-fuc / l-glu
SulfoSys - Biotec
No description specified

Contributor: Theresa Kouril

Assay type: Proteomics

Technology type: Tandem Mass Spectrometry

Snapshots: No snapshots

intracellular metabolome analysis: l-fucose vs d-glucose
SulfoSys - Biotec
No description specified

Contributor: Theresa Kouril

Assay type: Metabolite Profiling

Technology type: Gas Chromatography Mass Spectrometry

Snapshots: No snapshots

RNA sequencing:l-fuc/d-glu
SulfoSys - Biotec
No description specified

Contributor: Theresa Kouril

Assay type: RNA-seq Profiling

Technology type: Next generation sequencing

Snapshots: No snapshots

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Sso PGK in absence of recycling system, 31P NMR data
SulfoSys
No description specified

Creators: Theresa Kouril, Jacky Snoep

Contributors: Theresa Kouril, Jacky Snoep

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Experimental Data for Reconstituted Gluconeogenic System
SulfoSys

Experimental data for 3PG conversion to fructose-6-phosphase in reconstituted systems of gluconeogenesis of S. solfataricus

Creators: Jacky Snoep, Theresa Kouril

Contributor: Jacky Snoep

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Simulation for Reconstituted Gluconeogenic System
SulfoSys

Simulation results of experimental data of the reconstituted gluconeogenic system

Creators: Jacky Snoep, Theresa Kouril

Contributor: Jacky Snoep

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Temperature Degradation Kinetics Simulation
SulfoSys

Simulation results of temperature degradation of gluconeogenic intermediates

Creators: Jacky Snoep, Theresa Kouril

Contributor: Jacky Snoep

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TPI Kinetics Simulation
SulfoSys

Simulation results of TPI experimental data for GAP and DHAP saturation.

Creators: Jacky Snoep, Theresa Kouril

Contributor: Jacky Snoep

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GAPDH Kinetics Simulation
SulfoSys

Simulation results of GAPDH experimental data for BPG, NADPH, NADP, GAP, and Pi

Creators: Jacky Snoep, Theresa Kouril

Contributor: Jacky Snoep

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Sso PGK (28 ug/ml) in presence of recycling system, 31P NMR data
SulfoSys
No description specified

Creators: Theresa Kouril, Jacky Snoep

Contributor: Theresa Kouril

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Sso PGK (14 ug/ml) in presence of recycling system, 31P NMR data
SulfoSys
No description specified

Creator: Theresa Kouril

Contributor: Theresa Kouril

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PGK-GAPDH 30, 3PG, GAP & BPG
SulfoSys

incubations with yeast enzymes at 30C

Creators: Theresa Kouril, Jacky Snoep

Contributor: Theresa Kouril

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PGK-GAPDH 30, PYR
SulfoSys
No description specified

Creators: Theresa Kouril, Jacky Snoep

Contributor: Theresa Kouril

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PGK-GAPDH 70
SulfoSys
No description specified

Creators: Theresa Kouril, Jacky Snoep

Contributor: Theresa Kouril

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Sso PGK in presence of ATP recycling system at 70C, 3PG, BPG, PYR (enzymatic)
SulfoSys

metabolite quantification via enzymatic analysis

Creators: Theresa Kouril, Jacky Snoep

Contributor: Theresa Kouril

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BPG degradation at 70
SulfoSys
No description specified

Creators: Theresa Kouril, Jacky Snoep

Contributor: Theresa Kouril

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PGK yeast without recycling of ATP
SulfoSys, SulfoSys - Biotec

PGK yeast without recycling of ATP

Creators: Jacky Snoep, Theresa Kouril

Contributor: Jacky Snoep

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non-enzymatic phosphoenolpyruvate (PEP) degradation at 70°C and 80°C
SulfoSys - Biotec
No description specified

Creator: Theresa Kouril

Contributor: Theresa Kouril

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Activity assay L-fuc vs D-glc
SulfoSys - Biotec

Activity assays in cell free extracts of S. solfataricus grown on either L-fucose or D-glucose and activity assays with recombinant proteins

Creators: Jacqueline Wolf, Katharina Kruse, Bettina Siebers, Theresa Kouril

Contributor: Jacqueline Wolf

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Transcriptomics L-fuc vs D-glc
SulfoSys - Biotec

RNAseq analysis of S. solfataricus grown on either L-fucose or D-glucose

Creators: Jacqueline Wolf, Theresa Kouril, Andreas Albersmeier, Jörn Kalinowski

Contributor: Jacqueline Wolf

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RNA seq of l-Fucose/ d-Glucose
No description specified

Creator: Theresa Kouril

Contributor: Theresa Kouril

FBPAase Kinetic data
SulfoSys

Kinetic characterisation of FBPAase. Expermental data for enzyme reaction rates with increasing concentrations of DHAP and GAP.

Creators: Jacky Snoep, Theresa Kouril

Contributor: Jacky Snoep

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TPI Kinetic Data
SulfoSys

Kinetic characterisation of TPI. Experimental data for enzyme reaction rates with increasing concentrations of DHAP and GAP, and inhibition with 3PG and PEP.

Creators: Jacky Snoep, Theresa Kouril

Contributor: Jacky Snoep

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SULFOSYSbiotech_Proteome analysis
SulfoSys - Biotec

Protein extraction, iTRAQ labeling, peptides separation, mass spectrometry and data analysis

Creators: Theresa Kouril, Phil Wright, Trong Khoa Pham

Contributor: Theresa Kouril

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SulfoSYS_BIOTECH: RNA isolation and sequencing
SulfoSys - Biotec
No description specified

Creators: Theresa Kouril, Andreas Albersmeier, CeBiTech, University Bielefeld, Germany; Jörn Kalinowski, CeBiTech, University Bielefeld;

Contributor: Theresa Kouril

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SULFOSYSbiotech_Protein extraction_iTRAQ labeling_Peptides separation_mass spectrometry_data analysis
No description specified

Creator: Theresa Kouril

Contributor: Theresa Kouril

Preparation of cell extracts of recombinant E.coli
SulfoSys

Preparation of cell free extracts of the recombinant E. coli strains expressing the gluconeogenic S. solfataricus enzymes.

Creators: Jacky Snoep, Theresa Kouril

Contributor: Jacky Snoep

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Cloning and heterologous expression in E. coli
SulfoSys

Cloning and heterologous expression of gluconeogenic enzymes from S. solfataricus in E. coli

Creators: Jacky Snoep, Theresa Kouril

Contributor: Jacky Snoep

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Protein purification from E. coli extracts
SulfoSys

Purification of gluconeogenic enzymes from S. solfataricus in recombinant E.coli extracts

Creators: Jacky Snoep, Theresa Kouril

Contributor: Jacky Snoep

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The Carbon Switch at the Level of Pyruvate and Phosphoenolpyruvate in Sulfolobus solfataricus P2.
HOTSOLUTE

Abstract (Expand)

Sulfolobus solfataricus P2 grows on different carbohydrates as well as alcohols, peptides and amino acids. Carbohydrates such as D-glucose or D-galactose are degraded via the modified, branched … Entner-Doudoroff (ED) pathway whereas growth on peptides requires the Embden-Meyerhof-Parnas (EMP) pathway for gluconeogenesis. As for most hyperthermophilic Archaea an important control point is established at the level of triosephophate conversion, however, the regulation at the level of pyruvate/phosphoenolpyruvate conversion was not tackled so far. Here we describe the cloning, expression, purification and characterization of the pyruvate kinase (PK, SSO0981) and the phosphoenolpyruvate synthetase (PEPS, SSO0883) of Sul. solfataricus. The PK showed only catabolic activity [catalytic efficiency (PEP): 627.95 mM(-1)s(-1), 70 degrees C] with phosphoenolpyruvate as substrate and ADP as phosphate acceptor and was allosterically inhibited by ATP and isocitrate (K i 0.8 mM). The PEPS was reversible, however, exhibited preferred activity in the gluconeogenic direction [catalytic efficiency (pyruvate): 1.04 mM(-1)s(-1), 70 degrees C] and showed some inhibition by AMP and alpha-ketoglutarate. The gene SSO2829 annotated as PEPS/pyruvate:phosphate dikinase (PPDK) revealed neither PEPS nor PPDK activity. Our studies suggest that the energy charge of the cell as well as the availability of building blocks in the citric acid cycle and the carbon/nitrogen balance plays a major role in the Sul. solfataricus carbon switch. The comparison of regulatory features of well-studied hyperthermophilic Archaea reveals a close link and sophisticated coordination between the respective sugar kinases and the kinetic and regulatory properties of the enzymes at the level of PEP-pyruvate conversion.

Authors: Patrick Haferkamp, B. Tjaden, Lu Shen, C. Brasen, Theresa Kouril, Bettina Siebers

Date Published: 30th Apr 2019

Journal: Front Microbiol

Phosphoglycerate kinase acts as a futile cycle at high temperature.
SulfoSys, SulfoSys - Biotec

Abstract (Expand)

In (hyper)thermophilic organisms metabolic processes have to be adapted to function optimally at high temperature. We compared the gluconeogenic conversion of 3-phosphoglycerate via 1,3-bisphosphoglycerate … to glyceraldehyde-3-phosphate at 30 degrees C and at 70 degrees C. At 30 degrees C it was possible to produce 1,3-bisphosphoglycerate from 3-phosphoglycerate with phosphoglycerate kinase, but at 70 degrees C, 1,3-bisphosphoglycerate was dephosphorylated rapidly to 3-phosphoglycerate, effectively turning the phosphoglycerate kinase into a futile cycle. When phosphoglycerate kinase was incubated together with glyceraldehyde 3-phosphate dehydrogenase it was possible to convert 3-phosphoglycerate to glyceraldehyde 3-phosphate, both at 30 degrees C and at 70 degrees C, however, at 70 degrees C only low concentrations of product were observed due to thermal instability of glyceraldehyde 3-phosphate. Thus, thermolabile intermediates challenge central metabolic reactions and require special adaptation strategies for life at high temperature.

Authors: Theresa Kouril, J. J. Eicher, Bettina Siebers, Jacky Snoep

Date Published: 7th Oct 2017

Journal: Microbiology

Systems biology of the modified branched Entner-Doudoroff pathway in Sulfolobus solfataricus.
SulfoSys, SulfoSys - Biotec

Abstract (Expand)

Sulfolobus solfataricus is a thermoacidophilic Archaeon that thrives in terrestrial hot springs (solfatares) with optimal growth at 80 degrees C and pH 2-4. It catabolizes specific carbon sources, such … as D-glucose, to pyruvate via the modified Entner-Doudoroff (ED) pathway. This pathway has two parallel branches, the semi-phosphorylative and the non-phosphorylative. However, the strategy of S.solfataricus to endure in such an extreme environment in terms of robustness and adaptation is not yet completely understood. Here, we present the first dynamic mathematical model of the ED pathway parameterized with quantitative experimental data. These data consist of enzyme activities of the branched pathway at 70 degrees C and 80 degrees C and of metabolomics data at the same temperatures for the wild type and for a metabolic engineered knockout of the semi-phosphorylative branch. We use the validated model to address two questions: 1. Is this system more robust to perturbations at its optimal growth temperature? 2. Is the ED robust to deletion and perturbations? We employed a systems biology approach to answer these questions and to gain further knowledge on the emergent properties of this biological system. Specifically, we applied deterministic and stochastic approaches to study the sensitivity and robustness of the system, respectively. The mathematical model we present here, shows that: 1. Steady state metabolite concentrations of the ED pathway are consistently more robust to stochastic internal perturbations at 80 degrees C than at 70 degrees C; 2. These metabolite concentrations are highly robust when faced with the knockout of either branch. Connected with this observation, these two branches show different properties at the level of metabolite production and flux control. These new results reveal how enzyme kinetics and metabolomics synergizes with mathematical modelling to unveil new systemic properties of the ED pathway in S.solfataricus in terms of its adaptation and robustness.

Authors: Sofia Figueiredo, Theresa Kouril, D. Esser, Patrick Haferkamp, Patricia Wieloch, Dietmar Schomburg, Peter Ruoff, Bettina Siebers, Jörg Schaber

Date Published: 12th Jul 2017

Journal: PLoS One

A systems biology approach reveals major metabolic changes in the thermoacidophilic archaeon Sulfolobus solfataricus in response to the carbon source L-fucose versus D-glucose
SulfoSys - Biotec

Abstract (Expand)

Archaea are characterised by a complex metabolism with many unique enzymes that differ from their bacterial and eukaryotic counterparts. The thermoacidophilic archaeon Sulfolobus solfataricus is known … for its metabolic versatility and is able to utilize a great variety of different carbon sources. However, the underlying degradation pathways and their regulation are often unknown. In this work, we analyse growth on different carbon sources using an integrated systems biology approach. The comparison of growth on L-fucose and D-glucose allows first insights into the genome-wide changes in response to the two carbon sources and revealed a new pathway for L-fucose degradation in S. solfataricus. During growth on L-fucose we observed major changes in the central carbon metabolic network, as well as an increased activity of the glyoxylate bypass and the 3-hydroxypropionate/4-hydroxybutyrate cycle. Within the newly discovered pathway for L-fucose degradation the following key reactions were identified: (i) L-fucose oxidation to L-fuconate via a dehydrogenase, (ii) dehydration to 2-keto-3-deoxy-L-fuconate via dehydratase, (iii) 2-keto-3-deoxy-L-fuconate cleavage to pyruvate and L-lactaldehyde via aldolase and (iv) L-lactaldehyde conversion to L-lactate via aldehyde dehydrogenase. This pathway as well as L-fucose transport shows interesting overlaps to the D-arabinose pathway, representing another example for pathway promiscuity in Sulfolobus species. This article is protected by copyright. All rights reserved.

Authors: Jacqueline Wolf, Helge Stark, K. Fafenrot, Andreas Albersmeier, Trong Khoa Pham, K. B. Muller, B. Meyer, L. Hoffmann, L. Shen, Stefan Albaum, Theresa Kouril, K. Schmidt-Hohagen, M. Neumann-Schaal, C. Brasen, J. Kalinowski, Phil Wright, Sonja-Verena Albers, Dietmar Schomburg, Bettina Siebers

Date Published: 10th Sep 2016

Journal: Mol Microbiol

Intermediate instability at high temperature leads to low pathway efficiency for an in vitro reconstituted system of gluconeogenesis in Sulfolobus solfataricus
SulfoSys

Abstract (Expand)

Four enzymes of the gluconeogenic pathway in Sulfolobus solfataricus were purified and kinetically characterized. The enzymes were reconstituted in vitro to quantify the contribution of temperature … instability of the pathway intermediates to carbon loss from the system. The reconstituted system, consisting of phosphoglycerate kinase, glyceraldehyde 3-phosphate dehydrogenase, triose phosphate isomerase and the fructose 1,6-bisphosphate aldolase/ phosphatase maintained a constant consumption rate of 3-phosphoglycerate and production of fructose 6-phosphate over a 1 hour period. Cofactors ATP and NADPH were regenerated via pyruvate kinase and glucose dehydrogenase. A mathematical model was constructed on the basis of the kinetics of the purified enzymes and the measured half-life times of the pathway intermediates. The model quantitatively predicted the systems uxes and metabolite concentrations. Relative enzyme concentrations were chosen such that half the carbon in the system was lost due to degradation of the thermolabile intermediates dihydroxyacetone phosphate, glyceraldehyde 3-phosphate and 1,3 bisphosphoglycerate, indicating that intermediate instability at high temperature can significantly affect pathway efficiency. This article is protected by copyright. All rights reserved.

Authors: Theresa Kouril, Dominik Esser, Julia Kort, Hans Westerhoff, Bettina Siebers, Jacky Snoep

Date Published: 20th Jul 2013

Journal: FEBS J.

"Hot standards" for the thermoacidophilic archaeon Sulfolobus solfataricus
SulfoSys

Abstract (Expand)

Within the archaea, the thermoacidophilic crenarchaeote Sulfolobus solfataricus has become an important model organism for physiology and biochemistry, comparative and functional genomics, as well as, … more recently also for systems biology approaches. Within the Sulfolobus Systems Biology ("SulfoSYS")-project the effect of changing growth temperatures on a metabolic network is investigated at the systems level by integrating genomic, transcriptomic, proteomic, metabolomic and enzymatic information for production of a silicon cell-model. The network under investigation is the central carbohydrate metabolism. The generation of high-quality quantitative data, which is critical for the investigation of biological systems and the successful integration of the different datasets, derived for example from high-throughput approaches (e.g., transcriptome or proteome analyses), requires the application and compliance of uniform standard protocols, e.g., for growth and handling of the organism as well as the "-omics" approaches. Here, we report on the establishment and implementation of standard operating procedures for the different wet-lab and in silico techniques that are applied within the SulfoSYS-project and that we believe can be useful for future projects on Sulfolobus or (hyper)thermophiles in general. Beside established techniques, it includes new methodologies like strain surveillance, the improved identification of membrane proteins and the application of crenarchaeal metabolomics.

Authors: Melanie Zaparty, Dominik Esser, Susanne Gertig, Patrick Haferkamp, Theresa Kouril, Andrea Manica, Trong Khoa Pham, Julia Reimann, Kerstin Schreiber, Pawel Sierocinski, Daniela Teichmann, Marleen van Wolferen, Mathias Von Jan, Patricia Wieloch, Sonja-Verena Albers, Arnold Driessen, Hans-Peter Klenk, Christa Schleper, Dietmar Schomburg, John Van Der Oost, Phil Wright, Bettina Siebers

Date Published: 31st Aug 2009

Journal: Extremophiles

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