Nature Genetics
20, 46 - 50 (1998)
doi:10.1038/1700
Mass spectrometry and EST-database searching allows characterization
of the multi-protein spliceosome complexGitte Neubauer1, Angus King1, Juri Rappsilber1, Cinzia Calvio2, Mark Watson2, Paul Ajuh2, Judith Sleeman2, Angus Lamond2
& Matthias Mann1, 31
Protein & Peptide Group, European Molecular Biology
Laboratory, Meyerhofstr. 1, 69115
Heidelberg, Germany. 2
Department of Biochemistry, Dundee University,
Dundee DD1 4HN, United Kingdom. 3
Current address: Department of Molecular Biology, Odense
University, Campusvej 55, DK-5230
Odense M, Denmark.
Correspondence should be addressed to Matthias Mann mann@cebi.ou.dkMany important cell mechanisms are carried out and regulated by multi-protein
complexes, for example, transcription and RNA processing machinery, receptor
complexes and cytoskeletal structures. Most of these complexes remain only
partially characterized due to the difficulty of conventional protein analysis
methods. The rapid expansion of DNA sequence databases now provides whole
or partial gene sequences of model organisms, and recent advances in protein
microcharacterization via mass spectrometry allow the possibility of
linking these DNA sequences to the proteins in functional complexes1.
This approach has been demonstrated in organisms whose genomes have been sequenced2, such as budding yeast. Here we report the first characterization
of an entire mammalian multi-protein complex using these methods. The machinery
that removes introns from mRNA precursors — the spliceosome —
is a large multi-protein complex3,
4. Approximately half of
the components excised from a two-dimensional gel separation of the spliceosome
were found in protein sequence databases. Using nanoelectrospray mass spectrometry,
the remainder were identified and cloned using public expressed sequence tag
(EST) databases. Existing EST databases are thus already sufficiently complete
to allow rapid characterization of large mammalian protein complexes via
mass spectrometry.
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