A new crosslinker for mass spectrometric analysis of the quaternary structure of protein complexes

Mass spectrometric structural analysis of crosslinked peptides is a powerfu l method to elucidate the spatial arrangement of polypeptides in protein co mplexes. Our aim is to develop bifunctional crosslinkers that, after crossl inking protein complexes followed by proteolytic digestion, give rise to cr osslinked peptides that can be readily tracked down by mass spectrometry. T o this end we synthesized the crosslinker N-benzyliminodiacetoyloxysuccinim id (BID), which yields stable benzyl cation marker ions upon low-energy col lision-induced dissociation (CID) tandem mass spectrometry. Sensitive detec tion of the marker ion upon low-energy CID is demonstrated with different B ID-crosslinked peptide preparations. With BID it becomes possible to retrie ve crosslinked and crosslinker-adducted peptides, without the necessity of purifying crosslinked peptides prior to identification. The basic design of this crosslinker can be varied upon, in order to meet specific crosslinkin g needs, (C) 2001 American Society for Mass Spectrometry.