Many cellular processes are mediated by macromolecular assemblies that are too large or variable in structure to solve using crystallography and/or NMR. Moreover, crystallography and NMR require structures to be concentrated and removed from their biological milieu, therefore preventing analysis in the native state or in the context of other structures. TEM has fundamental limitations which include the need for thin samples, heavy metal stains, image averaging, and limited capabilities for detection of low atomic number elements. We propose to develop the Local Electrode Atom Probe (LEAP) to provide this information. With LEAP, sample atoms are removed one at a time using electrostatic ionization. Position and identity of removed atoms are determined by a position sensitive detector and time-of-flight mass spectrometry. LEAP has <0.5 nm positional resolution and near quantum efficiency, and therefore signal averaging is not required to obtain high resolution. We propose to develop sample preparation methods necessary for 3-D atomic-level resolution of biomacromolecules and macromolecular assemblies using the LEAP. Ultimately, we plan to extend this capability to provide 3-D atomic level structure of viruses and cells.
The LEAP has commercial applications in basic and clinical biology due to its ability to determine 3-D biological and biomolecular structures with higher resolution, far more complete elemental detail, and potentially more quickly than is possible with current imaging and analytical instruments such as SEMs and TEMs. In drug discovery, the LEAP can directly determine if candidate drugs bind with high avidity to target molecules, rather than to use molecular simulations as is currently done by the pharmaceutical industry, spending over $200M/yr. LEAP also has considerable applications in microelectronics, materials sciences, and biotechnology.
