The Biomolecular and Proteomics Mass Spectrometry Facility (BPMSF) at the University of California San Diego wishes to acquire a Synapt G2-S System with HXMS Automation. The PI of this project was the first to show that hydrogen/deuterium exchange (HX) could be measured by MALDI mass spectrometry. Her lab was one of the first to use this experimental approach for mapping macromolecular interfaces. Since 1998, the MALDI-TOF mass spectrometer in the BPMSF has been continuously used for HDX experiments, mainly performed by biochemists who were not experts in mass spectrometry. The Synapt G2-S System with HXMS Automation is the first fully-automated instrument that can replace the MALDI-TOF for use by non-mass spectrometry experts. The Waters instrument also provides many improvements in sensitivity, automation, and data analysis compared with the old manual MALDI-TOF approach. The integrated chromatographic system is based on the Waters nanoACQUITY platform and contains a Peltier-cooled module that houses the injection and switching valves, online pepsin digestion and C-18 separation columns. This chromatographic system is fully integrated with the qTOF mass spectrometer which combines dual-T-wave, off-axis ion transfer optics to transfer ions from the ion source to the quadrupole MS analyzer with the highest possible efficiency. A suite of software coordinates the sample preparation, chromatography, and mass spectrometry and provides rapid and automated data raw viewing, analysis and quantitation with the DynamX software. The Major Users are all researchers who have in the past, or will in the future attempt HDX experiments in the BPMSF. Their projects encompass the gamut of NIH-funded research. The Synapt G2-S System with HXMS Automation will enable new and important discoveries not possible with the outdated MALDI-TOF approach: Komives will measure the "foldedness" of inhibitors of NF- kappa B proteins and changes in their folded state upon binding to their targets;Ghosh will analyze conformational changes in the kinase (IKK) that phosphorylates these inhibitors targeting them for degradation;Noel will analyze conformational changes in Tobacco 5-epi-aristolochene synthase and other terpene synthases which they are engineering to make novel natural products;Gianneschi will probe the macrostructure of proteinaceous micelles for drug delivery;Jennings will analyze the conformational changes in interleukins and kinases;Shan will map the substrate- and receptor- interactions in the chloroplast Signal Recognition Particle;and Kay will screen modulators of circadian rhythm-control proteins.

Public Health Relevance

The Synapt G2-S System with HXMS Automation measures the degree of exposure of protein surfaces to solvent which gives vital information about the protein conformation and its interactions. Since the completion of the Human Genome Project, it has become clear that proteins have multiple forms depending on their modifications and interactions. To understand how specific gene products work to impact cellular function and human health, it is vital to understand the different functional forms of proteins.

Agency
National Institute of Health (NIH)
Institute
Office of The Director, National Institutes of Health (OD)
Type
Biomedical Research Support Shared Instrumentation Grants (S10)
Project #
1S10OD016234-01
Application #
8447332
Study Section
Special Emphasis Panel (ZRG1-BCMB-D (30))
Program Officer
Birken, Steven
Project Start
2013-09-01
Project End
2014-08-31
Budget Start
2013-09-01
Budget End
2014-08-31
Support Year
1
Fiscal Year
2013
Total Cost
$599,500
Indirect Cost
Name
University of California San Diego
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
804355790
City
La Jolla
State
CA
Country
United States
Zip Code
92093
Dembinski, Holly; Wismer, Kevin; Balasubramaniam, Deepa et al. (2014) Predicted disorder-to-order transition mutations in I?B? disrupt function. Phys Chem Chem Phys 16:6480-5