Mass spectrometry has become the enabling tool for identification of both lipids and proteins and for determination of their structures and properties. The changes in lipids with disease have become increasingly recognized, suggesting they may be biomarkers. One goal of the Washington University Mass Spectrometry Research Resource is to build a foundation for understanding lipid fragmentation, facilitating both de novo and computer-assisted identification of increasingly complex lipids found in decreasing amounts in bacteria and parasites. This work will continue because we recognize the growing challenge to identify more complex lipids and to uncover their role in infection and disease. In protein science and proteomics, related challenges exist to understand, with greater speed and sensitivity, protein properties and interactions. As a second goal, various chemical footprinting procedures that take advantage ofthe separation and analysis capabilities of modern chromatography and mass spectrometry will be explored as sources of information on protein structure, folding, and interfaces with metal ions, DNA, and other proteins. The strategy is to build a "tool box" of footprinting reagents that overtly modify proteins to map those regions that are accessible to reaction, thereby revealing interfaces and quantifying affinities. This approach takes full advantage of the power of modern "bottom-up" proteomics and is designed to be exportable to any proteomics laboratory. Nevertheless, bottom-up analytical proteomics fails when confronted with the need to identify the complex arrays of protein modifications that often form in post-translational modification or are introduced by footprinting. Addressing this problem by improving top-down proteomics is the third goal of the WU Resource. We propose to develop new instrumentation, methods, and associated data processing and interpretation strategies. They will be implemented along with a new high-field Fourier transform ion cyclotron resonance mass spectrometer to aid biomedical researchers with problems for which protein modification is an issue. Its success will serve to demonstrate a paradigm for top-down sequencing that can be employed by other researchers who search for the fine details of protein structure and function.

Public Health Relevance

Lipids and protein are important components in all living systems. Identifying them, characterizing their properties and interactions with other substances, and determining how they can be altered is important for understanding human health and disease. Mass spectrometry is meeting these challenges and moving forward with the innovative instrumentation and methods proposed here.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Biotechnology Resource Grants (P41)
Project #
8P41GM103422-35
Application #
8258753
Study Section
Special Emphasis Panel (ZRG1-BCMB-K (40))
Program Officer
Sheeley, Douglas
Project Start
1997-08-01
Project End
2014-12-31
Budget Start
2012-01-01
Budget End
2012-12-31
Support Year
35
Fiscal Year
2012
Total Cost
$1,385,601
Indirect Cost
$474,022
Name
Washington University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
Brennan, Patrick J; Tatituri, Raju V V; Heiss, Christian et al. (2014) Activation of iNKT cells by a distinct constituent of the endogenous glucosylceramide fraction. Proc Natl Acad Sci U S A 111:13433-8
McNulty, Samantha N; Fischer, Peter U; Townsend, R Reid et al. (2014) Systems biology studies of adult paragonimus lung flukes facilitate the identification of immunodominant parasite antigens. PLoS Negl Trop Dis 8:e3242
Wowor, Andy J; Yan, Yuetian; Auclair, Sarah M et al. (2014) Analysis of SecA dimerization in solution. Biochemistry 53:3248-60
Russler-Germain, David A; Spencer, David H; Young, Margaret A et al. (2014) The R882H DNMT3A mutation associated with AML dominantly inhibits wild-type DNMT3A by blocking its ability to form active tetramers. Cancer Cell 25:442-54
Zhang, Ying; Majumder, Erica L-W; Yue, Hai et al. (2014) Structural analysis of diheme cytochrome c by hydrogen-deuterium exchange mass spectrometry and homology modeling. Biochemistry 53:5619-30
Zhang, Hao; Cui, Weidong; Gross, Michael L (2014) Mass spectrometry for the biophysical characterization of therapeutic monoclonal antibodies. FEBS Lett 588:308-17
San Francisco, Brian; Sutherland, Molly C; Kranz, Robert G (2014) The CcmFH complex is the system I holocytochrome c synthetase: engineering cytochrome c maturation independent of CcmABCDE. Mol Microbiol 91:996-1008
Xu, Wei; Hsu, Fong-Fu; Baykal, Eda et al. (2014) Sterol biosynthesis is required for heat resistance but not extracellular survival in leishmania. PLoS Pathog 10:e1004427
Sattler, Fr; He, J; Chukwuneke, J et al. (2014) Testosterone Supplementation Improves Carbohydrate and Lipid Metabolism in Some Older Men with Abdominal Obesity. J Gerontol Geriatr Res 3:1000159
O'Sullivan, David; van der Windt, Gerritje J W; Huang, Stanley Ching-Cheng et al. (2014) Memory CD8(+) T cells use cell-intrinsic lipolysis to support the metabolic programming necessary for development. Immunity 41:75-88

Showing the most recent 10 out of 49 publications