| Autor: |
Waitt GM; Molecular Discovery Research, GlaxoSmithKline, Research Triangle Park, North Carolina 27709, USA., Xu R, Wisely GB, Williams JD |
| Jazyk: |
angličtina |
| Zdroj: |
Journal of the American Society for Mass Spectrometry [J Am Soc Mass Spectrom] 2008 Feb; Vol. 19 (2), pp. 239-45. Date of Electronic Publication: 2007 May 23. |
| DOI: |
10.1016/j.jasms.2007.05.008 |
| Abstrakt: |
Characterization of protein-ligand complexes by nondenaturing mass spectrometry provides direct evidence of drug-like molecules binding with potential therapeutic targets. Typically, protein-ligand complexes to be analyzed contain buffer salts, detergents, and other additives to enhance protein solubility, all of which make the sample unable to be analyzed directly by electrospray ionization mass spectrometry. This work describes an in-line gel-filtration method that has been automated and optimized. Automation was achieved using commercial HPLC equipment. Gel column parameters that were optimized include: column dimensions, flow rate, packing material type, particle size, and molecular weight cut-off. Under optimal conditions, desalted protein ions are detected 4 min after injection and the analysis is completed in 20 min. The gel column retains good performance even after >200 injections. A demonstration for using the in-line gel-filtration system is shown for monitoring the exchange of fatty acids from the pocket of a nuclear hormone receptor, peroxisome proliferator activator-delta (PPARdelta) with a tool compound. Additional utilities of in-line gel-filtration mass spectrometry system will also be discussed. |
| Databáze: |
MEDLINE |
| Externí odkaz: |
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