The soluble domains of Gpi8 and Gaa1， two subunits of glycosylphosphatidylinositol transamidase (GPI-T)， assemble into a complex

Glycosylphosphatidylinositol transamidase (GPI-T) catalyzes the post-translational addition of the GPI anchor to the C-terminus of some proteins. In most eukaryotes， Gpi8， the active site subunit of GPI-T， is part of a hetero-pentameric complex containing Gpi16， Gaa1， Gpi17， and Gab1. Gpi8， Gaa1， and Gpi16 co-purify as a heterotrimer from Saccharomyces cerevisiae， suggesting that they form the core of the GPI-T. Details about the assembly and organization of these subunits have been slow to emerge. We have previously shown that the soluble domain of S. cerevisiae Gpi8 (Gpi8) assembles as a homodimer， similar to the caspases with which it shares weak sequence homology (Meitzler， J. L. et al.， 2007). Here we present the characterization of a complex between the soluble domains of Gpi8 and Gaa1. The complex between GST-Gpi8 (α) and His-Gaa1 (β) was characterized by native gel analysis and size exclusion chromatography (SEC) and results are most consistent with an αβ stoichiometry. These results demonstrate that Gpi8 and Gaa1 interact specifically without a requirement for other subunits， bring us closer to determining the stoichiometry of the core subunits of GPI-T， and lend further credence to the hypothesis that these three subunits assemble into a dimer of a trimer.