Structural and functional characterizations of the proteasome-activating protein PA26 from Trypanosoma brucei

The activated 20 S proteasome, which has been found only in mammalian cells , is composed of two heptamer rings of an activator protein on each end of the 20 S proteasome and is inducible by interferon-gamma, A 20 S proteasome has been recently identified in a protozoan pathogen Trypanosoma brucei, b ut there has been no experimental evidence yet for the presence of a 26 S p roteasome. Instead, an activated form of 20 S proteasome was isolated from this organism, which has significantly enhanced peptidase activities. It co nsists of an additional activator protein with an estimated molecular mass of 26 kDa (PA26) (To, W, Y,, and Wang, C, C, (1997) FEES Lett, 404, 253-262 ), The profile and sequences of tryptic peptides from PA26 were determined by mass spectrometry; no matches were found in the data base. The peptide s equences were used in reverse transcriptase-polymerase chain reaction to is olate a full-length cDNA clone encoding PA26, The protein sequence thus der ived from it indicates little sequence identity with those of mammalian act ivator proteins PA28 alpha, beta, or gamma, There is only a single copy of PA26 gene in T, brucei, Purified recombinant PA26 polymerizes spontaneously to form heptamer ring with an outer diameter of 8.5 nm, The ring binds and activates 20 S proteasomes from T, brucei as well as rat, whereas human PA 28 alpha can neither bind nor activate T, brucei 20 S proteasome. The forme r is thus apparently more ubiquitous than PA28 in its capability of binding to and activating 20 S proteasomes. Its presence in T, brucei may also sug gest a more ancient origin of proteasome activator proteins and a much wide r involvement in protein degradation among other eukaryotic organisms than was originally envisaged.