Proteins that co-purify with tandem affinity-purified PRC5 in Trypanosoma brucei

The project was to determine the role of a trypanosomatid protein, termed PRC5, in pre-mRNA splicing. PRC5 has no homologs in model organisms but was recurrently co-purified with known splicing factors in Trypanosoma brucei. To determine interacting proteins, PRC5 was C-terminally TAP-tagged (we used the PTP tag) and tandem affinity-purified, and co-purifying proteins identified after separation by SDS-PAGE and in-gel trypsin digest by LC/MS/MS. Many of the enriched proteins were known splicing factors, and sucrose gradient sedimentation identified a complex of five subunits, termed PRP19-related complex or PRC.