Proteomics Core

Robert Cole   Director
Robert Cole, PhD
rcole@jhmi.edu
410-614-6968
  Mark Donowitz   Associate Director
Mark Donowitz, MD
mdonowit@jhmi.edu
410-955-9675

The Hopkins Digestive Diseases Basic and Translational Research Core Center (HDDBTRCC) Proteomics Core uses mass spectrometry coupled to one (1D) and two (2D) dimensional separations by column chromatography or gel electrophoresis to identify, quantify or characterize proteins and their post-translational modifications, that are expressed in well characterized protein fractions from the small intestine, colon, kidney, liver and pancreas. Techniques such as difference gel electrophoresis (DIGE), isobaric tag for relative and absolute quantitation (iTRAQ), tandem mass tags (TMT) and stable isotope labeling of amino acids in cell culture (SILAC) as well as non-labeling methods (MudPIT, multi-dimensional protein identification technology) are available for quantifying relative differences in protein expression and post-translational modifications, such as acetylation, glycosylation, phosphorylation, nitrosation, ubiquitination and novel cleavage sites. Also available are Core developed methods to characterize post-translational modifications, such as LCMS methods to accurately determine the mass of intact proteins, a selective fluorescent labeling of cysteines to detect oxidized or nitrosated cysteines, or enrichment strategies to map acetylation, phosphorylation and ubiquitin sites. This Core is a part of larger institutional resource and provides a Proteomics Specialist to assist Center investigators and their staff in preparing and analyzing their samples or in using the Core’s multi-user equipment. Through the Proteomics Specialist, the Center investigators have access to all the staff and expertise in the Johns Hopkins School of Medicine Mass Spectrometry and Proteomics Facility.

Goals 
The specific goals of this Core are to help Center Investigators identify and quantify differentially expressed proteins in normal or diseases of the small intestine, colon, kidney, liver and pancreas, track interactions with binding partners during changes in signal transduction, identify proteolytic cleavage sites and map post-translational modifications.

Equipment

    Q-Exactive Orbitrap (Thermo)
    LTQ Orbitrap Velos (Thermo)
    Voyager DE-STR MALDI-TOF (AB Sciex)
    Typhoon 9400

Services
- Consultation, pre- and post-analysis
- Sample preparation: Buffer exchange; Column Chromatography; Proteolytic digestion; Stable isotope labeling
- Protein Identification of proteins in solution, in gel bands or spots, or in complex protein mixtures
- Protein Modifications: acetylation, AMPylation, citrullination, glycosylation, phosphorylation, proline hydroxylation, nitrosation, ubiquitination and novel cleavage sites
- Protein Quantification: Isobaric Tagging for Relative and Absolute Quantitation (iTRAQ), Tandem Mass Tags (TMT), Absolute Quantification (AQUA), Protein Standard Absolute Quantification (PSAQ).
- High Resolution Mass Analysis of Intact Proteins: protein characterization and their modifications
- Biostatical/Bioinformatic analysis: PCA analysis, Volcano plots, Ingenuity based gene ontology pathway analysis; bioinformatics consultation.
- Training Workshops: MALDI, 2D-Gel Electrophoresis
- Self Service: MALDI, 2D-Gel Electrophoreis, Scanners, Bioinformatic Software


Mass Spectrometry and Proteomic Facility