The P2 protein of peripheral nervous system myelin induces experimental allergic neuritis in rats, a model of Guillain-Barre syndrome in humans. Previous purification procedures have used acid extraction to obtain the protein in lipid-free form (LF-P2). Here, we have purified the P2 protein in lipid-bound form (LB-P2) by extracting myelin with the detergent CHAPS, followed by Cu2+-affinity column chromatography. All myelin lipids were present in the preparation as shown by high-performance thin-layer chromatography and mass spectrometry. The LB-P2 preparation, which differs from LF-P2 in solubility and in the secondary-structure composition, was dialyzed to remove unbound lipids and excess detergent and crystallized using the hanging-drop vapor diffusion technique. Crystals of lipid-bound P2 appeared usually very reproducibly within 2 weeks at pH 5.7 in polyethylene glycol 6000 (PEG6000) at concentrations of 20-30% (w/v), and larger crystals were obtained by additional sitting-drop crystallization. Xray diffraction showed reflections up to 2.7 A. The crystallization conditions (25-30% PEG6000, pH 5.0) and the unit cell dimensions (a = 94.5 Angstrom, b = 94.5 Angstrom, c = 74.2 Angstrom, a 90degrees, gamma = 120degrees) of LB-P2 were different from those earlier described for LF-P2 (10% PEG4000, pH 3, and unit cell dimensions a = 91.8 Angstrom, b = 99.5 Angstrom, c = 56.5 Angstrom, alpha = beta = gamma = 90.0degrees). It is important that P2 has been crystallized with specifically bound lipids; therefore, solving this new crystal structure will reveal details of this protein's behavior and role in the myelin sheath. (C) 2003 Elsevier Science (USA). All rights reserved.