The National Deuteration Facility (NDF*) has developed expertise in biological molecular deuteration for the in vivo deuteration of proteins, where all or a fraction of the hydrogen atoms in an expressed protein are replaced by the stable isotope deuterium (²H).
The NDF capability for the production of multiple-labelled proteins using ¹³C and ¹⁵N is of particular interest to the NMR community for both solution and solid state NMR applications. As with deuterium (²H), the isotopes ¹³C and/or ¹⁵N are introduced into the minimal growth medium utilised for biomass production and protein expression using the NDF methods for high-yield recombinant expression of partial or per-deuterated protein in Escherichia coli. Selective-labelling of desired amino acids with ¹³C and/or ¹⁵N in recombinantly expressed proteins has now also been demonstrated by the NDF with robust methods continuing to be developed and refined. The fungal hydrophobin EASdelta15, previously triple-labelled (²H/¹³C/¹⁵N) for solid state NMR analysis in order to investigate the molecular structure of the fibrillar form, has been selectively labelled with ¹³C/¹⁵N asparagine and phenylalanine for continuing structural investigations. Along with an overview of NDF labelling methods for uniform and selective labelling, examples of protein deuteration and multiple labelling will be highlighted for NMR and neutron instrument investigations.
The NDF offers facilities, staff and expertise to the neutron beam and NMR user communities through an externally refereed proposal scheme, accessible at http://www.ansto.gov.au/ndf