Identification and isolation of three proteasome subunits and their encoding genes from Trypanosoma brucei

We have determined peptide sequences of three Trypanosoma brucei proteasome subunit proteins by mass spectrometry of tryptic digests of the proteins p urified by two-dimensional (2-D) polyacrylamide gel electrophoresis. Three genes identified by the sequence of their cDNA encode the peptides identifi ed in these three proteins. The three proteins predicted from the gene sequ ences have significant similarity to other known proteasome subunits and re present an alpha 6 type subunit (TbPSA6), and two beta-type subunits belong ing to the beta 1-type (TbPSB1) and beta 2 type (TbPSB2). The sequences of both beta-subunits predict formation of catalytically active subunits throu gh proteolytic processing. The prediction is supported by the presence in e ach of the two beta-subunits of a tryptic peptide that has the correctly pr ocessed N-terminus that creates the threonine nucleophile of the mature pro tein. This peptide cannot be generated by trypsin because of the required c leavage of a glycine-threonine bond. It is thus likely that there are at le ast two catalytically active beta-subunits, TbPSB1 and TbPSB2, present in t he mature 20S proteasome from T. brucei. (C) 1999 Elsevier Science B.V. All rights reserved.