The addition of glycosylphosphatidylinositol (GPI) anchors to eukaryotic proteins in the lumen of the endoplasmic reticulum is catalyzed by the transamidase complex, composed of at least five subunits (PIG-K, PIG-S, PIG-T, PIG-U and GPAA1). Here PIG-K24-337 and PIG-S38-467 from yeast, including the residues 24-337 and 38-467 of the entire 411 and 534 residue protein, respectively, was produced in Escherichia coli and purified to homogeneity. Analysis of secondary structure by circular dichroism spectroscopy showed that yPIG-K24-377 comprises 52% alpha-helix and 12% beta-sheet, whereas yPIG-S38-467 involves 58% alpha-helix and 18% beta-sheet. The radius of gyration (R-g) and the maximum size (D-max) of both proteins have been analyzed by small angle X-ray scattering (SAXS) and determined to be 2.64 +/-0.3 and 10.3 +/- 0.1 nm (yPIG-K24-377) as well as 3.06 +/- 0.02 nm (R-g) and 16.9 +/- 0.4 nm (D-max) in the case of yPIG-S38-467, respectively. Using an ab initio approach, the first low-resolution solution structures of both proteins were restored. yPIG-K24-377 is an elongated particle consisting of an egg-like portion and a small globular segment linked together by an 1.9 nm long stalk. yPIG-S38-467 forms an elongated molecule in solution with a larger domain of 10.1 nm in length, a diameter of 9.1 nm and a smaller domain of 6.7 nm in length and 3.4 nm in width. The two domains of yPIG-S38-467 are tilted relative to each other. Finally, the arrangements of PIG-K and PIG-S inside the ensemble of the transamidase complex are discussed. (C) 2010 Elsevier Inc. All rights reserved.