Tetramerization of glycosylphosphatidylinositol-specific phospholipase C from Trypanosoma brucei

Glycosylphosphatidylinositol-specific phospholipase C (GPI-PLC) is an integ ral membrane protein in the protozoan parasite Trypanosoma brucei, Enzyme a ctivity appears to be suppressed in T, brucei, although the polypeptide is readily detectable. The basis for the apparent quiescence of GPI-PLC is not known. Protein oligomerization was investigated as a possible mechanism fo r post-translational regulation of GPI-PLC activity, An equilibrium between monomers, dimers, and tetramers of purified GPI-PLC was detected by molecu lar sieving and shown to be perturbed with specific detergents. Homotetrame rs dominated in Nonidet P-40, and dimers and monomers of GPI-PLC were the m ajor species in 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate. The detergents were exploited as tools to study the effect of oligomerizati on on enzyme activity. Tetrameric GPI-PLC was 3.6-20-fold more active than the monomeric enzyme. Tetramer existence was confirmed by chemical cross-li nking. In vivo cross-linking revealed the oligomeric state of GPI-PLC durin g latency and after enzyme activation. During quiescence, monomers were the predominant species in T, brucei, Assembly of tetrameric GPI-PLC occurred when parasites were subjected to conditions known to activate the enzyme. I n Leishmania where heterologous expression of GPI-PLC causes a GPI deficien cy, the enzyme existed as a tetramer, Hence, oligomerization of GPI-PLC is associated with high enzyme activity both in vivo and in vitro.