Developing graphene grids to overcome air-water-interface issues
Ended Project Grant Awarded Last action
- Award ID
- R21GM154216
- Recipient
-
Scripps Research Institute UEI PHZJFZ32NKH4
- Awarding agency
-
National Institute of General Medical Sciences
National Institutes of Health, Department of Health and Human Services
- Obligated
- $506,000
Award facts
- Assistance listing
- 93.859 Biomedical Research and Research Training
- Completion date
- Funding agency
- Assistant Secretary for Financial Resources Department of Health and Human Services
- Place of performance
- La Jolla, CA 92037, USA
Award details come from USAspending.gov.
Description
- Abstract
- Single particle cryoem is now a ubiquitously employed methodology for high-resolution structure determination of biomedically important macromolecular complexes. Cryoem structures have revealed the detailed mechanisms underlying a wide range of cellular functions and helped understand how environmental or genetic factors perturb biological function to give rise to disease. However, all biological specimens prepared for single particle cryoem imaging are exposed to the hydrophobic air-water interface during the sample preparation process. This interaction with the air-water interface has a destructive effect on the structural integrity of the targeted specimen, resulting in partial or complete unfolding of proteins, compositional dissociation of complexes, and widespread aggregation of the sample. This interaction with the hydrophobic air-water interface poses the single largest challenge facing the field of single particle cryoem, and severely limits the utility of cryoem structure determination for a wide range of samples. Particularly stable samples that maintain structural integrity after repeated interactions with the air-water interface often adopt preferred orientations relative to the hydrophobic surface, which either diminishes or completely eliminates one's ability to determine a high-resolution structure of the targeted specimen. While there are additives and newer sample preparation devices that aim to mitigate these issues arising from the air-water interface, none of these are broadly applicable for all samples, nor do they fully overcome the air-water interface issues. Thus, cryoem researchers often spend as much time (or more) optimizing cryoem sample preparation methods as in establishing expression and purification conditions for a given macromolecule. Given that all commonly used cryoem sample preparation technologies expose particles to the air-water interface, most of the acquired particle data are damaged and must be computationally discarded during image analysis. This makes cryoem structure determination incredibly inefficient. We propose to overhaul the grid preparation process to abolish air-water interface interactions during cryoem grid prep to accommodate preservation of fragile, hard-to-purify samples while substantially improving the efficiency of cryoem data collection. This new technology will minimize damage to protein samples by blocking interaction with the air-water interface, which will increase applicability and efficacy of cryo-EM data collection, while also enabling robust quantification of macromolecular abundance or conformational dynamics present in a sample. Using a combination of graphene and nanocage technologies we will develop a cheap and robust methodology for cryoem sample preparation that abolishes air-water interface interactions. Successful execution of the proposed sample preparation platform will advance our understanding of macromolecular mechanisms by enabling the preservation of challenging samples that thus far been recalcitrant to single particle cryoem studies. This innovative cryoem grid system will enable researchers to probe unexplored aspects of cell biology, and have a transformative impact that will reverberate throughout the structural biology community.
Transactions
| Action | Obligated | Date |
|---|---|---|
| Amendment | $230,000 | |
| Original award | $276,000 |
Quick answers
Who received award R21GM154216?
R21GM154216 was awarded to Scripps Research Institute by National Institute of General Medical Sciences (National Institutes of Health) on September 15, 2024.
How much is award R21GM154216 worth?
$506,000 has been obligated on R21GM154216.
Which agency awarded R21GM154216?
National Institute of General Medical Sciences (National Institutes of Health) awarded R21GM154216, with funding from Assistant Secretary for Financial Resources (Department of Health and Human Services).
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