This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.Dysregulation of proteolytic processing of the amyloid precursor protein (APP) contributes to the pathogenesis of Alzheimer's Disease, and the Group VIA phospholipase A2 (iPLA2B) is the dominant PLA2 enzyme in the central nervous system and is subject to regulatory proteolytic processing. We have identified novel N-terminal variants of iPLA2B and previously unrecognized proteolysis sites in APP constructs with a C-terminal 6-myc tag by automated identification of signature peptides in LC/MS/MS analyses of proteolytic digests. We have developed a Signature-Discovery (SD) program to characterize protein isoforms by identifying signature peptides that arise from proteolytic processing in vivo. This program analyzes MS/MS data from LC analyses of proteolytic digests of protein mixtures that can include incompletely resolved components in biological samples. This reduces requirements for purification and thereby minimizes artifactual modifications during sample processing. A new algorithm to generate the theoretical signature peptide set and to calculate similarity scores between predicted and observed mass spectra has been tested and optimized with model proteins. The program has been applied to the identification of variants of proteins of biological interest, including APP cleavage products and iPLA2B, and such applications demonstrate the utility of this approach.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
5P41RR000954-30
Application #
7597575
Study Section
Special Emphasis Panel (ZRG1-BPC-H (40))
Project Start
2007-02-01
Project End
2008-01-31
Budget Start
2007-02-01
Budget End
2008-01-31
Support Year
30
Fiscal Year
2007
Total Cost
$1,130
Indirect Cost
Name
Washington University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
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