DETERMINATION OF PROTEIN FOLD RECOGNITION USING CHEMICAL CROSS-LINKING AND MASS SPECTROMETRY (SF99-065)

Determination of the three-dimensional structures of proteins has traditionally been achieved by x-ray crystallography and NMR. These techniques produce high resolution structural data but have various limitations including requirement for large amount of pure protein and extensive length of time for data analysis. There are also several purely computational methods for predicting protein fold, but these methods are generally unreliable and have low resolution.

UCSF investigators have developed a fast and efficient technique for determining the 3D structure or conformation of proteins using a combination of chemical cross-linking and analytical strategies, mass spectrometry, and software analysis. The technique is practiced in stages, beginning with the identification of the fundamental fold pattern of a protein domain, followed by placement of the domains with respect to each other. Finally the 3D structure can be obtained within 3-6 Å or better depending on the constraints and modeling approach employed. One key advantage this technique has over other methods is that the cross-linking reaction is performed best at low concentration (<10 ?M) and the chemistry can accommodate the presence of other reagents or ligands that might be required for the protein to exhibit its native structure or conformation.

ADVANTAGES:

• Readily applied to other biological macromolecules.



• Can be practiced on macromolecules in non-crystalline environments (eg. in aqueous solution, in cytoplasm, on membrane-bound components).



• Cost effective:
  • Allows rapid data acquisition and analysis.
  
  
  • Requires low amount of protein (0.1-1 mg).

REFERENCES:
Young, M.M., et al. High throughput protein fold identification by using experimental constraints derived from intramolecular cross-links and mass spectrometry. (2000) Proc. Nat’l. Acad. Sciences 97: 5802.

Schilling, , et al. MS2Assign, automated assignment and nomenclature of tandem mass spectra of chemically crosslinked peptides (2003) J. Amer. Soc. Mass Spectrom. 14(8): 834.

INTELLECTUAL PROPERTY: The Regents of the University of California have filed a patent application covering methods of determining the 3D conformation of a protein.
PCT Application title and number: Method of Determining the Three-Dimension of a Macromolecule, WO2000US0014667.
(http://www.delphion.com/details?pn=WO00072004A3)

If you would like to receive further information about this technology and potential licensing opportunities, please contact:

Sunita Rajdev, Ph.D.
Licensing Officer
Phone: (415) 353-4470
Fax: (415) 348-1579
Sunita.rajdev@ucsf.edu

Reference: OTM Case #SF99-065