The surface of Trypanosoma brucei brucei insect forms is covered by an invariant protein coat consisting of procyclins. There are six or seven procyclin genes that encode unusual proteins with extensive tandem repeat units of glutamic acid (E) and proline (P) (referred to as EP repeats), and two genes that encode proteins with internal pentapeptide (GPEET) repeats. Although the EP forms of procyclins have been isolated and characterized by several laboratories, evidence for GPEET procyclin has largely been confined to the expression of its mRNA. To characterize GPEET procyclin further, we isolated the protein from T. b. brucei strain 427. We found that label from [3H]myristic acid and [3H]ethanolamine was incorporated into GPEET procyclin and we demonstrated the protein's covalent modification with a glycosylphosphatidylinositol anchor. The major form of GPEET procyclin showed an apparent molecular mass of 22–32 kDa, was susceptible to proteolytic treatment and was found to be phosphorylated. Surprisingly, our results show that GPEET procyclin represents the major form of procyclin in T. b. brucei 427 culture forms and that the ratio of EP to GPEET procyclin can vary considerably between different cell lines.

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