High throughput screening using acoustic droplet ejection to combine protein crystals and chemical libraries on crystallization plates at high density
Abstract
Here, we describe a high throughput method for screening up to 1728 distinct chemicals with protein crystals on a single microplate. Acoustic droplet ejection (ADE) was used to co-position 2.5 nL of protein, precipitant, and chemicals on a MiTeGen in situ-1 crystallization plate™ for screening by co-crystallization or soaking. ADE-transferred droplets follow a precise trajectory which allows all components to be transferred through small apertures in the microplate lid. The apertures were large enough for 2.5 nL droplets to pass through them, but small enough so that they did not disrupt the internal environment created by the mother liquor. Using this system, thermolysin and trypsin crystals were efficiently screened for binding to a heavy-metal mini-library. Fluorescence and X-ray diffraction were used to confirm that each chemical in the heavy-metal library was correctly paired with the intended protein crystal. A fragment mini-library was screened to observe two known lysozyme ligands using both co-crystallization and soaking. A similar approach was used to identify multiple, novel thaumatin binding sites for ascorbic acid. This technology pushes towards a faster, automated, and more flexible strategy for high throughput screening of chemical libraries (such as fragment libraries) using as little as 2.5 nL of each component.
- Authors:
- Publication Date:
- Research Org.:
- Brookhaven National Laboratory (BNL), Upton, NY (United States)
- Sponsoring Org.:
- USDOE Office of Science (SC), Office of Workforce Development for Teachers & Scientists (WDTS); USDOE Office of Science (SC), Biological and Environmental Research (BER); USDOE Office of Science (SC), Basic Energy Sciences (BES); USDOE Office of Science (SC), Workforce Development for Teachers and Scientists (WDTS)
- OSTI Identifier:
- 1250356
- Alternate Identifier(s):
- OSTI ID: 1229106; OSTI ID: 1341502
- Report Number(s):
- BNL-111181-2015-JA; BNL-108331-2015-JA
Journal ID: ISSN 1047-8477; S1047847715300034; PII: S1047847715300034
- Grant/Contract Number:
- 11-008; AC02-98CH10886; SC0012704; SC00112704
- Resource Type:
- Published Article
- Journal Name:
- Journal of Structural Biology
- Additional Journal Information:
- Journal Name: Journal of Structural Biology Journal Volume: 191 Journal Issue: 1; Journal ID: ISSN 1047-8477
- Publisher:
- Elsevier
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 36 MATERIALS SCIENCE; high throughput screening; fragment screening; drug discovery; chemical biology; acoustic droplet ejection; in situ x-ray data collection; crystallography; synchrotron radiation
Citation Formats
Teplitsky, Ella, Joshi, Karan, Ericson, Daniel L., Scalia, Alexander, Mullen, Jeffrey D., Sweet, Robert M., and Soares, Alexei S. High throughput screening using acoustic droplet ejection to combine protein crystals and chemical libraries on crystallization plates at high density. United States: N. p., 2015.
Web. doi:10.1016/j.jsb.2015.05.006.
Teplitsky, Ella, Joshi, Karan, Ericson, Daniel L., Scalia, Alexander, Mullen, Jeffrey D., Sweet, Robert M., & Soares, Alexei S. High throughput screening using acoustic droplet ejection to combine protein crystals and chemical libraries on crystallization plates at high density. United States. https://doi.org/10.1016/j.jsb.2015.05.006
Teplitsky, Ella, Joshi, Karan, Ericson, Daniel L., Scalia, Alexander, Mullen, Jeffrey D., Sweet, Robert M., and Soares, Alexei S. Wed .
"High throughput screening using acoustic droplet ejection to combine protein crystals and chemical libraries on crystallization plates at high density". United States. https://doi.org/10.1016/j.jsb.2015.05.006.
@article{osti_1250356,
title = {High throughput screening using acoustic droplet ejection to combine protein crystals and chemical libraries on crystallization plates at high density},
author = {Teplitsky, Ella and Joshi, Karan and Ericson, Daniel L. and Scalia, Alexander and Mullen, Jeffrey D. and Sweet, Robert M. and Soares, Alexei S.},
abstractNote = {Here, we describe a high throughput method for screening up to 1728 distinct chemicals with protein crystals on a single microplate. Acoustic droplet ejection (ADE) was used to co-position 2.5 nL of protein, precipitant, and chemicals on a MiTeGen in situ-1 crystallization plate™ for screening by co-crystallization or soaking. ADE-transferred droplets follow a precise trajectory which allows all components to be transferred through small apertures in the microplate lid. The apertures were large enough for 2.5 nL droplets to pass through them, but small enough so that they did not disrupt the internal environment created by the mother liquor. Using this system, thermolysin and trypsin crystals were efficiently screened for binding to a heavy-metal mini-library. Fluorescence and X-ray diffraction were used to confirm that each chemical in the heavy-metal library was correctly paired with the intended protein crystal. A fragment mini-library was screened to observe two known lysozyme ligands using both co-crystallization and soaking. A similar approach was used to identify multiple, novel thaumatin binding sites for ascorbic acid. This technology pushes towards a faster, automated, and more flexible strategy for high throughput screening of chemical libraries (such as fragment libraries) using as little as 2.5 nL of each component.},
doi = {10.1016/j.jsb.2015.05.006},
journal = {Journal of Structural Biology},
number = 1,
volume = 191,
place = {United States},
year = {Wed Jul 01 00:00:00 EDT 2015},
month = {Wed Jul 01 00:00:00 EDT 2015}
}
https://doi.org/10.1016/j.jsb.2015.05.006
Web of Science
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