Silantes Lab Tour

Silantes produces stable isotope labelled biomolecules for NMR and mass spectrometry applications. All products are made using a cascade of biotechnology fermentations without any chemical synthesis steps. Utilizing a variety of fermenters, Silantes produces cell growth media, ready to use NMR standards, labelled nucleotides, phosphoramidites, amino acids and many other biomolecules.

StabIe isotope labelled biomolecules are used for applications in NMR and mass spectrometry. SI labelled amino acids are in particularly high demand for quantifying and analysing proteins by mass spectrometry.

In the fermentation shown in Fig. 1, Ralstonia eutropha (Cupriavidus necator) is the host strain of choice due to its facultative chemolithoautotrophic metabolism, which indicates that the organism is able to change its metabolic pathway according to the substrate available.

While both autotrophic and heterotrophic fermentations are possible, the latter is considerably more expensive due to the required organic carbon sources.

The metabolic end product of this fermentation is cyanophycin, which forms the base for the production of the stable isotope labeled amino acids arginine and aspartate.

16 liter fermenter used for culturing Ralstonia eutropha bacteria.
Figure 1. 16 liter fermenter used  for culturing Ralstonia eutropha bacteria.

SI labelled amino acids are of particular interest for quantitative proteomics applications with arginine used for the SILAC method in cell culture. A major advantage of this method is the fact that cells growing in the presence of labelled amino acids incorporate the label into their proteins without any perturbing effects on their metabolism

Silantes 13C labelling workflow benefits

Ralstonia eutropha is fermented in the presence of 13CO2 and incorporates the label in all components of its metabolic pathways. One metabolic product is cyanophicin, which is an intermediate for the production of the amino acids arginine and aspartic acid.

With regard to the production of 13C-labelled substances, autotrophic organisms that utilize 13CO2 as their sole carbon source, such as algea or Ralstonia eutropha, are very cost efficient. In addition to the fact that 13CO2 is cheaper than 13C-labelled sugars, heterotrophs in comparison to autotrophs incorporate only a fraction of the label, with the rest lost due to respiration.

Article: https://journals.asm.org/doi/full/10.1128/AEM.01972-12

Zero waste principle

Once the Ralstonia eutropha fermentation is completed, the SI labelled biomass is introduced as an organic carbon source in a second cascade step of another heterotrophic fermentation to harvest additional complex biomolecules.

Green Compact

Silantes is also using green algae as another autotrophic production organism that utilizes photosynthesis to generate organic material with water, CO2 and light. Plate or tube reactors are most commonly used for these applications. Silantes has designed the “Green compact” with incorporated glass rods that support the fermentation with illumination from the inside (Fig. 2-5). Compared to other fermenters the “Green compact” has an optimal space time yield, improved ergonomics,and increased robustness and stability.

Microalgae, in this case Chlorella vulgaris, have the ability to convert inorganic into organic compounds. When they are cultured in the presence of 13CO2,15NO3– or 2H2O their biomass is labelled with the respective stable isotope, and a variety of stable isotope-labelled compounds can be extracted and purified.

Silantes Green Compact fermentor before innoculation
Fig. 2. Silantes Green Compact fermenter before innoculation
Green compact fermentor 2H labelled Chlorella vulgaris 2 days after inoculation
Fig. 3. 2H labelled Chlorella vulgaris 2 days after inoculation
2H labelled Chlorella vulgaris 1 week after inoculation
Fig. 4. 2H labelled Chlorella vulgaris 1 week after inoculation
Green Compact Fermentor with 2H labelled Chlorella vulgaris 2 weeks after inoculation
Fig. 5. 2H labelled Chlorella vulgaris 2 week after inoculation

Article: https://www.sciencedirect.com/science/article/abs/pii/0304416570900929

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