Capture and analysis of low molecular weight ligands by surface plasmon resonance combined with mass spectrometry

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Eur. J. Mass Spectrom. 7, 385 - 391 (2001)

Capture and analysis of low molecular weight ligands by surface plasmon resonance combined with mass spectrometry

Carsten P. Sünksen and Peter Roepstorff^*
Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense University, Campusvej 55, DK-5230 Odense, Denmark
Per-Olof Markgren and U. Helena Danielson
Department of Biochemistry, Uppsala University, Biomedical Center, S-751 23 Uppsala, Sweden
Markku D. Hämäläinen and Östen Jansson
Biacore AB, S-754 50 Uppsala, Sweden

ABSTRACT:

The combination of biomolecular interaction analysis (BIA) by surface plasmon resonance (SPR) and nano-electrospray ionization ion trap mass spectrometry (nanoESI-Ion Trap MS) as well as matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-ToF MS) is demonstrated for the binding of low molecular weight inhibitors (~ 600 Da) to HIV-1 protease. Inhibitors were captured on sensor chips of a manual or an automated SPR biosensor, to which HIV-1 protease was immobilized. Compounds and buffer components that bound unspecifically to the sensor surface were removed and the inhibitors were eluted in a minimal volume (3 ŠL), between air bubbles, in order to prevent dispersion of analyte into buffer eluent. Molecular weights were subsequently determined by mass spectrometry, structural information was obtained by MALDI-ToF post-source decay as well as by electrospray ionization tandem mass spectrometry (MS/MS) analysis. Furthermore, competition experiments, using a mixture of different ligands, demonstrated that the peak intensities in the MALDI-ToF spectrum could be used for relative quantification of the amount of the different ligands bound to the immobilized target. Methodology for automated capture and elution of analytes was developed and evaluated.

Keywords: surface plasmon resonance, matrix-assisted laser desorption/ionization, electrospray ionization, HIV-1 protease inhibitors

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