hide
Free keywords:
Laser induced liquid bead ion desorption-mass spectrometry (LILBID-MS); F1Fo-ATP synthase; RNA polymerase; Membrane protein complexes; Proteomics
Abstract:
Mass spectrometry of large macromolecules is still a methodological challenge. We here report on the
application of the recently developed LILBID (laser induced liquid bead ion desorption) mass spectrometry
by which the biomolecules dissolved in microdroplets are desorbed/ablated by a mid-IR laser into vacuum.
Two modes of desorption are possible: an ultrasoft mode at low laser intensity in which a macromolecule is
desorbed as integral complex into vacuum and a harsher mode at higher intensity, by which it is dissociated
into its covalent subunits. With this method we studied the soluble core polymerase II and a membrane-
embedded F1Fo-ATP synthase, solubilized by detergent. For both complexes the complete complex in
different charge state is observed at ultrasoft conditions. At elevated laser intensities all 10 subunits could
be assigned for the core Pol II. In the case of the ATP synthase under equal conditions all eight subunits
appear in the mass spectrum, assigned by a correspondence of the expected theoretical masses of the
subunits and the observed ones. In addition the method requires only sample volumes of microliter at
micromolar concentration and is tolerant to detergents. Therefore it is a low consumptive method well
adapted for the mass analysis of biomolecules of low availability such as membrane molecules.
