? The proposed research involves characterization of large protein complexes such as vault proteins using mass spectrometry (MS) and proteomics to elucidate structure and biological function. Vaults are linked to multidrug resistance (MDR), the major cause for chemotherapy failure in cancer treatment. Although the function of vaults remains unknown, several functions are proposed such as a cellular transporter or in signaling pathways for cell survival or growth. Identification of vault interacting proteins and vault contents using MS, immunoprecipitation, chemical cross-linking, and/or tandem affinity purification (TAP) tagging of the major vault protein (MVP) will lead toward the elucidation of its interacting components, structural assembly, and biological function. This will also confirm the role of vaults in MDR if drugs are found in vault derived MDR established cell lines. Since MVP is a novel substrate of a protein tyrosine phosphatase (SHP-2), the phoshorylation sites of MVP will be identified by 2D-PAGE, and stained for phosphoproteins, trypsin digested, enriched by affinity chromatography or an automated method by a titanium oxide column, and analyzed by HPLC-ESI-MS/MS, only requiring femtomolar levels of phosphopeptides. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Predoctoral Individual National Research Service Award (F31)
Project #
5F31GM075384-04
Application #
7495524
Study Section
Special Emphasis Panel (ZRG1-F08 (29))
Program Officer
Toliver, Adolphus
Project Start
2005-09-06
Project End
2010-09-05
Budget Start
2008-09-06
Budget End
2009-09-05
Support Year
4
Fiscal Year
2008
Total Cost
$29,587
Indirect Cost
Name
University of California Los Angeles
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
092530369
City
Los Angeles
State
CA
Country
United States
Zip Code
90095
Lomeli, Shirley H; Peng, Ivory X; Yin, Sheng et al. (2010) New reagents for increasing ESI multiple charging of proteins and protein complexes. J Am Soc Mass Spectrom 21:127-31
Lomeli, Shirley H; Yin, Sheng; Ogorzalek Loo, Rachel R et al. (2009) Increasing charge while preserving noncovalent protein complexes for ESI-MS. J Am Soc Mass Spectrom 20:593-6
Kaddis, Catherine S; Lomeli, Shirley H; Yin, Sheng et al. (2007) Sizing large proteins and protein complexes by electrospray ionization mass spectrometry and ion mobility. J Am Soc Mass Spectrom 18:1206-16