Use of stable isotope labeled growth media for protein and lipid NMR analyses
In the following article, we present two publications from the journal Nature Communications. In both papers stable isotope labeled rich growth media were used for NMR protein interaction analysis. The media were produced for culturing E. coli. As shown in the second paper, the media can also be used for other bacteria
How did stable isotope protein expression advance the research on human norovirus?
Human norovirus infection is the most common cause of gastroenteritis worldwide. However, due to the frequent occurrence of new virus variants, antivirals and vaccines have made treatment and prevention challenging.
The 2019 Nature Communications paper entitled “A post-translational modification of human Norovirus capsid protein attenuates glycan binding” sheds light on the mechanism of infection. Stable-isotope-labeled-rich-growth-media were used.
Binding of human noroviruses to histo blood group antigens (HBGAs) is relevant for the infection, but how this step promotes the infection of host cells is unknown. Using protein- nuclear magnetic resonance, mass spectrometry and crystallography, researchers investigated HBGA binding to the P- domain of a viral strain.
What is the result and application of E. coli OD2 DN medium?
Post-translational modifications of asparagine were identified that have effects on HBGA- detection and therefore indicate relevance in infections & immune recognition of many noro strains.
P-domain asparagine post-translational modifications were identified to affect HBGA binding and may therefore be of relevance for noro strain infections & immune recognition.
Source: Mallagaray, A., Creutznacher, R., Dülfer, J. et al. A post-translational modification of human Norovirus capsid protein attenuates glycan binding. Nat Commun 10, 1320 (2019). https://doi.org/10.1038/s41467-019-09251-5
Click for more Information about the product used: OD2 Media solution for E.coli
How they are able to support the analysis of the antibiotic teixobactin?
The antibiotic teixobactin is used for many clinically relevant multidrug-resistant bacteria. So far, no resistance to the antibiotic has been identified and the development of resistance to teixobactin is difficult. Its mechanism of action needs to be explored in order to develop analogues with improved properties.
The binding of teixobactins to cellular membranes which is involved in its mode of action was studied in a 2020 paper entitled ”Mode of action of teixobactins in cellular membranes” using Silantes stable isotope rich media.
Results and Application of E.coli OD2 CN medium
It was found that the teixobactin binding affinity for lipid ll in cell membranes is weakened under certain conditions, which suggests that the direct interaction of the antibiotic with cell wall precursors is not the only mechanism involved in killing bacteria.
Silantes [13C,15N]-labeled rich medium was used to grow S.simulans 22. This allowed the generation of substrates for the synthesis of lipid ll by enzymatic lipid reconstitution.
Source: Shukla, R., Medeiros-Silva, J., Parmar, A. et al. Mode of action of teixobactins in cellular membranes. Nat Commun 11, 2848 (2020).
What makes stable isotope labeled growth media a valuable tool for structural analysis by NMR spectroscopy?
Structural studies on proteins using nuclear magnetic resonance (NMR) spectroscopy require labelling with stable isotopes. This can be achieved by expression of the protein of interest in bacteria and yeast. The above papers provide practical insight into stable isotope labelling and protein expression for NMR studies.
The papers discussed in this article are licensed under a Creative Commons Attribution 4.0 International. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/