Protein analysis by mass spectrometry and sequence database searching: Tools for cancer research in the post‐genomic era

The post‐genomic era is characterized by the deposition of sequence information for entire genomes in databases. Currently, besides the protein sequences for known human proteins, there are partial sequences from thousands more human proteins for which no biological function has been assigned. A powerful new tool for the unambiguous identification and characterization of gel‐separated proteins is accomplished by the combination of mass spectrometry and sequence database searching. This combination provides the cancer biologist with the ability to (i) identify the potential protein:protein associations and (ii) fully characterize function‐critical post‐translational modifications, both directly from silver‐stained polyacrylamide gels. In this report we describe the application of tandem mass spectrometry and database searching to two problems which are prototypical for cancer research and indeed for biomedical research in general. The first is the identification of gel‐separated, low abundance proteins based on amino acid sequence composition following coimmunoprecipitation with the human apoptosis inhibitor protein BclXL. The second is the determination of the precise sites of phosphorylation of the human regulatory protein 4E‐BP1, which controls mRNA translation.

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