SulfoSys - Biotec

e:Bio - Innovations Competition Systems Biology

Present in many industrial effluents and as intermediate of lignin degradation, phenol is a widespread pollutant causing serious environmental problems, due to its toxicity to animals and humans. Removal of phenol from the environment by bacteria has been studied extensively over the past decades, but only little is known about phenol biodegradation in hostile environments. We combined metabolomics and transcriptomics together with metabolic modelling to elucidate the organism’s response to growth ...

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

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 ...

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

To investigate phenol degradation in Saccharolobus solfataricus transcriptome and metabolome analyses were performed with cells grown on phenol as sole carbon source. Cells grown on D-glucose served as reference. Metabolic modelling was used to compare efficiency of phenol utilization in terms of oxygen demand and energy yield with the reference condition.

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.

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

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

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

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

Genome scale metabolic model of Sulfolobus solfataricus specific scenario: modelling of L-fucose degradation pathways

Intracellular and extracellular metabolome analysis

Growth on D-glucose served as reference scenario

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

A model for the PGK reaction of yeast in presence or absence of the ATP recycling reactions

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

BPG degradation at 70C

BPG stability analysis

PGK 70C model

PGK - GAPDH models

Reference genome of Sulfolobus solfataricus P2 used for mapping of RNA-seq results. Based on NCBI genome of Sulfolobus solfataricus.

RNA-Seq raw data: Sulfolobus solfataricus P2 grown on casaminoacids and d-glucose.

RNA-Seq mapping results (part1): Sulfolobus solfataricus P2 grown on casaminoacids and d-glucose, mapped using bowtie2 against reference sequence of Sulfolobus solfataricus P2 (NBCI, see other data files of experiment).

RNA-Seq mapping results (part2): Sulfolobus solfataricus P2 grown on casaminoacids and d-glucose, mapped using bowtie2 against reference sequence of Sulfolobus solfataricus P2 (NBCI, see other data files of experiment).

PGK yeast without recycling of ATP

PGK yeast with recycling of ATP

Supplementary file required by main Zip archive file for file extraction.

Cells of S. solfataricus were grown on either caseinhydrolysate or D-glucose (reference) as sole carbon source.

The distribution of enzymes involved in oxidative Stickland reactions among archaea was estimated using BLAST searches (BLOSUM62) with the protein sequences of acetate-CoA ligase (EC 6.2.1.13, ACS), ketoisovalerate oxidoreductase (EC 1.2.7.7, BC-OR) and indolepyruvate oxdoreductase (EC 1.2.7.8, AR-OR) from Pyrococcus furiosus (Pfu) and Sulfolobus solfataricus (Sso) against all archaea. Positive results are indicated by a '+' (homologue found, e-value < 1e-20) and negative results by a '-' (no ...

The calculation of amino acid uptake rates for cells of Sulfolobus solfataricus P2 grown on caseinhydrolysate was performed based on the relative consumption of individual amino acids from the growth medium and the previously published absolute concentration of amino acids in the used growth medium.

Protein, RNA and DNA content during the logarithmic growth phase were investigated in this study. The values represent the average of at least three independent experiments. Errors represent the standard deviation between the experiments.

Sulfolobus solfataricus P2 was cultivated on 1 % Caseinhydrolysate. Samples of the culture supernatants were taken at regular intervalls and analyzed by GC-MS. To evaluate the stability of amino acids under cultivation conditions an additional non-inoculated control culture was also analyzed.

Intracellular metabolome analysis of S. solfataricus P2 grown on caseinhydrolysate or D-glucose as sole carbon source. Samples were analyzed with GC-MS. CoA derivatives were analyzed with LC-MS.

These files contain the output results from Phenyx searching, providing all necessary information for matching MS data with protein database.

This file contains the results from proteome analysis of Sulfolobus solfataricus grown on Caseinhydrolysate as carbon source (growth on D-glucose served as reference condition).

Raw MS data files of the comparison of Sulfolobus solfataricus grown on either caseinhydrolysate or D-glucose. The numbers represents the fractions collected from the HPLC run (e.g. 22 indicates that this sample was collected from 21 min to 22 min). For MS analysis every two fractions were combined, indicated by double numbers (e.g. 80-81: combined fractions 80 and 81). Furhter all combined fractions were run twice on the MS. Zip archive. Requires the supplementary .z01, .z02 and .z03 files for ...

BPG stability notebook

PGK model for S. solfataricus

PGK-GAPDH model Sulfolobus kouril8

PGK-GAPDH model yeast kouril7

PGK-GAPDH models yeast and Sulfolobus Fig. 4 in manuscript

PGK 70C SBML

PGK yeast Fig1a

PGK yeast with/without recycling

Metabolic model of Sulfolobus solfataricus P2 in the SBML (sbml) and metano (txt, sce, fba) format. Scenarios are specific for growth on D-glucose or caseinhydrolysate as sole carbon source.

Metabolic model of Sulfolobus solfataricus P2 in the SBML (xml) and metano (txt, sce, fba) format. Scenarios are specific for growth on D-glucose or L-fucose as sole carbon source. Different theoretical routes of L-fucose degradation were modeled (E. coli-like, Xanthomonas-like and lactaldehyde-forming). Highest overall agreement between the model and experimental data was observed for the lactaldehyde-forming route.

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