Thursday, November 21, 2024
11/21/2024
The structures of biomacromolecules at atomic resolution (< 2.0-2.5 Å) are of enormous importance to understand their physiological functions and roles in diseases. An exemplary critical need of high atomic resolution is to resolve the location of proton/hydrogen which plays vital roles in various biological processes. Deuteration renders neutron scattering techniques unique advantages in high contrast (signal/background) to locate D/H. Like X-ray crystallography which has contributed majority of known biomolecule structures, high quality single crystals are the prerequisites for both X-ray and neutron data collection. It is worth mentioning that despite the recent progresses in electron microscopy techniques, true atomic resolution remains a formidable challenge to achieve. Lower resolution structures are associated with ambiguity and could mislead basic biomedical research as well as drug design/development applications. With the understanding on the fundamental limitations and technical hurdles associated with currently adopted ensemble-based methods, we propose to develop a single-entity method (named NanoAC) which will offer unprecedented capability to synthesize crystals one at a time, under real-time monitoring and with predictive crystal quality. A single nanotip will be employed to spatially confine supersaturation as the sole nucleation site. Electroanalytical and optical methods will monitor the whole crystallization process in real-time to capture quantitative signatures for the nucleation and crystal growth at single entity resolutions. Those signatures will enable active controls in kinetic transitions, and be quantitatively correlated with its diffraction quality and/or crystal habits. The insights will inform crystal synthesis such that nucleation kinetics and growth rates of each individual crystal will be finetune to improve crystal quality and to tune crystal size/habits. Prototype soluble proteins, nucleic acids and membrane proteins will be used as defined in this early-stage technology development program. The new toolbox, once established, will provide paradigm-shift capabilities to improve the crystal quality in diffraction and size/habit controls, to tackle challenging material systems currently not-crystallizable, and also feature high efficiency in time and/or materials. The overarching goal will be pursued through three interrelated aims. Aim 1 will establish real-time monitoring signatures for the generalization of NanoAC to crystallize soluble biomacromolecules and complexes. Aim 2 will correlate diffraction quality and crystal habits with monitoring signatures. Aim 3 will further develop single nanopipettes as ‘magic wand’ to crystallize membrane proteins.
HHS’ Tracking Accountability in Government Grants System (TAGGS) website provides access to detailed descriptions of grants, loans, aggregated direct payments and other types of financial assistance awarded by the HHS Operating Divisions (OPDIVs) and the Office of the Secretary Staff Divisions (STAFFDIVs).
