EM 3D contour maps provide protein assembly at the nanoscale within the neuronal porosome complex

The neuronal porosome complex, the secretory machinery at the plasma membrane of nerve terminals, is a 12-17-nm cup-shaped lipoprotein structure possessing a central plug. Since the porosome is a membrane associated, multi-protein complex measuring >650 kD, it has precluded generation of 3D crystals for x-ray diffraction studies, nor structural analysis at the atomic level using solution magnetic resonance spectroscopy. These limitations were partially overcome in the current studies, furthering our understanding of the porosome structure. Using atomic force microscopy, electron microscopy and electron density and 3D contour mapping, finally provides at the nanoscale, the structure and assembly of proteins within the neuronal porosome complex. Results from this study demonstrate a set of eight protein units lining the porosome cup, each connected via spoke-like elements to a central plug region within the structure. The isolation of intact porosomes for near-atomic resolution using cryo-electron diffraction measurements, is finally possible.