Mass spectrometry and EST-database searching allows characterization of the multi-protein spliceosome complex

Gitte Neubauer, Angus King, Juri Rappsilber, Cinzia Calvio, Mark Watson, Paul Ajuh, Judith Sleeman, Angus Lamond, Matthias Mann (Lead / Corresponding author)

    Research output: Contribution to journalArticlepeer-review

    407 Citations (Scopus)

    Abstract

    Many 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 complexes. This approach has been demonstrated in organisms whose genomes have been sequenced, 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 complex. 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.
    Original languageEnglish
    Pages (from-to)46-50
    Number of pages5
    JournalNature Genetics
    Volume20
    Issue number1
    DOIs
    Publication statusPublished - Sep 1998

    Keywords

    • Amino Acid Sequence
    • Base Sequence
    • DNA, Complementary
    • Databases, Factual
    • Electrophoresis, Gel, Two-Dimensional
    • Gene Expression
    • Green Fluorescent Proteins
    • Humans
    • Luminescent Proteins
    • Macromolecular Substances
    • Mass Spectrometry
    • Molecular Sequence Data
    • Proteins
    • Recombinant Proteins
    • Sequence Homology
    • Spliceosomes

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