Characterization of the <i>In Vivo</i> Deuteration of Native Phospholipids by Mass Spectrometry Yields Guidelines for Their Regiospecific Customization

Customization of deuterated biomolecules
is vital for many advanced
biological experiments including neutron scattering. However, because
it is challenging to control the proportion and regiospecificity of
deuterium incorporation in live systems, often only two or three synthetic
lipids are mixed together to form simplistic model membranes. This
limits the applicability and biological accuracy of the results generated
with these synthetic membranes. Despite some limited prior examination
of deuterating Escherichia coli lipids in vivo, this approach has not been widely implemented.
Here, an extensive mass spectrometry-based profiling of E. coli phospholipid deuteration states with several
different growth media was performed, and a computational method to
describe deuterium distributions with a one-number summary is introduced.
The deuteration states of 36 lipid species were quantitatively profiled
in 15 different growth conditions, and tandem mass spectrometry was
used to reveal deuterium localization. Regressions were employed to
enable the prediction of lipid deuteration for untested conditions.
Small-angle neutron scattering was performed on select deuterated
lipid samples, which validated the deuteration states calculated from
the mass spectral data. Based on these experiments, guidelines for
the design of specifically deuterated phospholipids are described.
This unlocks even greater capabilities from neutron-based techniques,
enabling experiments that were formerly impossible.