Mass Spectrometry Facility

	Principal Investigator: Dr. Michael Goshe Phone:919-513-7740 Office Location: Polk Hall 43 michael_goshe@ncsu.edu
	MS Laboratory Supervisor: Mr. Kevin Blackburn Phone: 919-515-0833 Office Location: Polk Hall 49 kevin_blackburn@ncsu.edu

Lab Location: Polk Hall 42
Lab Phone: 919-515-0831

Biological Mass Spectrometry Overview

Mass spectrometry is a powerful technique used frequently for both qualitative and quantitative characterization of proteins and other biomolecules as well as their interactions. In a typical MS experiment, molecules are ionized, accelerated into a mass analyzer, separated on the basis of mass-to-charge ratio (m/z), and detected. In another type of experiment known as MS/MS, one or more "precursor" ions are fragmented inside a collision cell in the mass spectrometer, and the resulting "product" ions mass analyzed. The resulting product ion spectrum may be used to provide insights into molecular structure based on the pattern of product ions generated. In combination with other techniques such as liquid chromatography, gel electrophoresis, chemical crosslinking, and co-immunoprecipitation, a variety of complex biological problems may be addressed.

Research

The mass spectrometry facility in Molecular and Structural Biochemistry is used to support a variety of research efforts through collaboration with Dr. Goshe. Although not a fee-for-service core facility, a large number of collaborative efforts are supported for faculty across departments and colleges, including generation of preliminary data in support of planned grant submissions on a case by case basis. Ongoing efforts can be broadly categorized as follows.

Proteomics: Characterization of the composition and dynamics of proteomes in plant and animal systems using a variety of approaches including label-free quantitation, SILIP, SILAC, and other custom labeling techniques. Determination of protein interaction partners through the use of co-immunoprecipitaton and other affinity based techniques.
Characterization of Post-Translational Modifications: Site assignment and quantitative characterization of post-translational modifications including phosphorylation, ubiquitination, and others. Site-specific stoichiometry measurements in response to various stimuli or environmental conditions may be determined.
Protein Structure: Through the use of a variety of novel chemical crosslinking reagents, protein-protein interactions are studied. Insights into protein structure and folding may be obtained through the analysis of individual proteins following chemical crosslinking. Similarly, intramolecular disulfide bonding patterns within proteins may be determined by MS. In addition, quantitative compositional analysis of protein macromolecular complexes often provides insights into the structure and function of the complex or individual components.

Facilities

The mass spectrometry laboratory located in the basement of Polk Hall is comprised of approximately 1100 square feet of customized space including electrical, ventilation, and specialty gases for mass spectrometry equipment. Two additional wet labs of approximately 900 square feet located on the 1st floor of Polk Hall are used for sample prep.

Instrumentation

	Waters NanoAcquity UPLC and Q-Tof Premier UPLC (Ultra-Performance Liquid Chromatograph) for nanoscale LC separations at up to 10,000 psi. Q-Tof Premier MS uses ESI for LC/MS, LC/MS/MS, or LC/MS^E analyses. MALDI ionization is also available on this instrument. Maximum mass resolution is 17,500 FWHM in W-mode, with mass errors of < 5 ppm with nanolockspray.
	Waters CapLC and Q-Tof API-US CapLC for nanoscale or capillary LC separations. Q-Tof API-US MS uses ESI for LC/MS or LC/MS/MS analyses. Infusion studies are also conducted on this instrument. Maximum mass resolution is 10,000 FWHM in W-mode.
	Thermo Finnigan TSQ-Quantum Triple quadrupole MS capable of a variety of MS/MS experiments including product ion scanning, precursor ion scanning, selected reaction monitoring (SRM), and multiple reaction monitoring (MRM).
	Agilent 1100 CapLC and Finnigan LCQ Deca (2 systems) Agilent 1100 for high-resolution capillary LC separations coupled to a Finnigan LCQ Deca ion trap MS via a custom nanospray ion source. These instruments are capable of multi-stage MS/MS experiments (MSⁿ) providing unique capabilities for characterization of PTMs such as phosphorylation and glycosylation. An additional LCQ Deca XP is also available.

Outreach

September, 2009 — Scouts from Boy Scout Troop 55 and Cub Scout Pack 91 recently visited the Mass Spectrometry and X-ray Crystallography Facilities. The boys toured both laboratories and participated in a variety of hands-on activities and experiments demonstrating how chemistry is involved in their everyday lives. For their participation, the Boy Scouts earned their Chemistry Merit Badge while the Cub Scout earned his Science Belt Loop.

Useful Links

Professional Associations

Triangle Area Mass Spectrometry Discussion Group (http://biochem.ncsu.edu/TAMS/)
American Society for Mass Spectrometry (http://asms.org)
Association for Biomolecular Resource Facilities (http://www.abrf.org)
Human Proteome Organization (http://www.hupo.org)