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SYMB: Massenspektrometrie in der Bioanalytik
SYMB 1: Massenspektrometrie in der Bioanalytik
SYMB 1.2: Hauptvortrag
Freitag, 17. März 2006, 11:00–11:30, HV
Biomoleküle - Laserdesoriert aus Mikrotröpfchen — •Bernd Brutschy — Institut für Physikalische und Theoretische Chemie, JW Goethe-Universität, Marie-Curie-Str. 9-11, 60439 Frankfurt
Mass spectrometry (MS) of biomolecules was revolutionized by electrospray ionization MS (ESI) and matrix-assisted laser desorption/ ionization MS (MALDI), honored by the Nobel Prize in 2002. Both methods have specific advantages and draw-backs and are now widely applied in biology, biochemistry and pharmacology. Since some years an alternative laserdesorption method is being developed in our group called LILBID-MS (laser induced liquid beam/bead/blob ion desorption)[1], which tries to combine some of the advantages of the above methods.
Here ions are desorbed/ablated from micro droplets (φ=50 µ m V=65 pl) into vacuum by resonant excitation of vibrations in the solvent by means of a ns-pulsed IR OPO-laser (∼ 3 µ m ).
LILBID proved to be a highly sensitive and soft method which is relatively tolerant to salt and buffers. It allows the investigation of biomolecules in native solution at very low analyte consumption, typically at femto- to attomolar (10-18 ) or even lower amount. The duty cycle of the method is 1. Not only noncovalently bonded biomolecules are detectable at relatively low charge state, but also specific noncovalently bonded biomolecular complexes [2]. For their specifity (molecular recognition) most often a specific native environment (buffer, pH, solvent) is mandatory, which can be realized in the droplets. The method is applicable to a wide range of biomolecules from smaller peptides and aminoacids, to isolated RNA and DNA. The latter may be targeted by specific ligands (antibiotics and inhibitors). But also large hydrophobically bound membrane proteins such as cytochrome-C-oxidase (COX 125 kD) may be detected. A further advantage of this methods is that it allows to test the binding strenght of a complex qualitatively by varying the intensity of the laser. Thus by thermolysis, larger complexes like COX can be systematicly fractionalized into its molecular subunits.
The talk will give an overview of the experiment, discuss some results and presents a possible mechanism for the ion formation.
[1] W. Kleinekofort, J. Avdiev, B. Brutschy: A new method of laser desorption mass spectrometry for the study of biological macromolecules Int. J. Mass. Spectr. Ion Proc. 152 (1996) 135-142
[2] N. Morgner, H.-D. Barth, B. Brutschy: A new way to detect noncovalently bonded complexes of biomolecules from liquid micro-droplets by laser mass spectrometry accepted for publication in Australian Journal of Chemistry, Volume59, No 1 (2006)