InsPecT: Fast and accurate identification of post-translationally modified peptides from tandem mass spectra

Reliable identification of post-translational modifications is key to understanding various cellular regulatory processes. We describe a tool, InsPecT, to identify post-translational modifications using tandem mass spectrometry data. InsPecT constructs database filters that proved to be very successful in genomics searches. Given an MS/MS spectrum S and a database D, a database filter selects a small fraction of database D that is guaranteed (with high probability) to contain a peptide that produced S. InsPecT uses peptide sequence tags as efficient filters that reduce the size of the database by a few orders of magnitude while retaining the correct peptide with very high probability. In addition to filtering, InsPecT also uses novel algorithms for scoring and validating in the presence of modifications, without explicit enumeration of all variants. Inspect identifies modified and modified peptides with better or equivalent accuracy than other database search tools while being two orders of magnitude faster than SEQUEST, and substantially faster than X!TANDEM on complex mixtures. The tool was used to identify a number of novel modifications in different data-sets, including many phosphopetides in data provided by Alliance for Cellular Signalling that were missed by other tools.

The interaction of SV40 small tumor antigen with protein phosphatase 2A stimulates the map kinase pathway and induces cell proliferation

Interaction with SV40 small tumor antigen (small t) compromised the ability of multimeric protein phosphatase 2A to inactivate the mitogen-activated protein kinase ERK1 and the mitogen-activated protein kinase kinase MEK1. Transient expression of small t in CV-1 cells activated MEK and ERK but did not affect Raf activity. Small t stimulated the growth of quiescent CV-1 cells almost as effectively as did serum. Coexpression of kinase-deficient ERK2 blocked most, but not all, of the proliferation caused by small t. Activation of the mitogen-activated protein kinase pathway and stimulation of cell growth were dependent on the interaction of small t with protein phosphatase 2A. These findings indicate that SV40 small t is capable of inducing cell growth through blockade of protein phosphatase and deregulation of the mitogen-activated protein kinase cascade