Peptides and Proteomics Core
Equipment & Services
Proteomics Core
The Proteomics Core occupies approximately 2,000 square feet at the University of Michigan. Major equipment at the University of Michigan site includes:
- two Applied Biosystems model 4700 MALDI TOF/TOF mass spectrometers for proteome mapping, identification of post-translational modifications, and iTRAQ quantitation with 10 ppm mass accuracy and sensitivity to one femtomole
- LCQ Deca with capillary HPLC for LCMSMS and 2D LCMSMS with 200 ppm mass accuracy
- Waters MALDI QTOF for proteome mapping and analysis of post-translational modifications with 10 ppm mass accuracy
- Applied Biosystems DE PRO MALDI TOF for peptide mass fingerprinting, development, and QC
Agilent HPLC with MALDI fraction collector for interfacing LC with the TOFTOF
- Eldex capillary HPLC for microscale SCX
- Gilson preparative HPLC for large scale SCX separations (phosphoproteome applications)
- three spot-picking and sample handling robots to support 2D gels
- laser and transmittance scanners for imaging 2D gels
- 2D gel apparati with capacity for up to 40 gels per day
Additional equipment available through the Michigan Proteome Consortium at the 1,000 sq. ft. spoke laboratory at Michigan State University includes an LCQ XP with electrospray and MALDI interfaces, Micromass QTOF Ultima, and a Thermo-Finnegan LTQ FTMS. The MSU site is not supported by this application but these resources are available to investigators as needed. A photographic darkroom, environmental rooms, autoclave, and meeting room are all in close proximity to the University of Michigan Proteome Core. Excellent instrument and electronics shops are located in the Chemistry and Physics departments and available to the laboratory. The Proteomics Core is adjacent to the National Resource for Proteomics and Pathways (NRPP) and shares some resources and personnel including the Director, Dr. Andrews. The NRPP UM laboratory occupies approximately 800 square feet (rooms 1188 and 1195 of NIB) of wet laboratory space and includes an ultracentrifuge, high-speed preparative centrifuge, IEF and 2D Gel equipment, gel scanner, cell culture equipment, HPLC, PCR, apparatus for organic synthesis, and other typical laboratory equipment.
The Proteomics Core has a complete IT infrastructure and a full-time systems administrator to manage several Linux and Windows servers, and several PC and Solaris workstations. The NRPP and the Michigan Proteome Consortium share use of several of the servers. The laboratory maintains software for image analysis and an extensive system for information management. The Proteome Consortium also operates a 4 cpu NT server, a linux cluster, and several NT servers for administrative and database search tools. The Consortium has site licenses for Mascot, SeQuest, Protein Prospector, and Protein Global Server and maintains an X!Tandem (3) server and an eight cpu Linux cluster. Additional Linux cluster support will be provided by the Michigan Center for Biological Information as needed.
Protein Structure Core
The Protein Core currently occupies 3,900 square feet of modern laboratory space and offices. The Core is located in contiguous space on the same floor of Medical Science Research Building II, is well equipped for complete protein structure analysis and peptide synthesis, and is in close proximity to the other BRCF laboratories.
Major equipment in the Protein Structure Core includes:
- The Symphony Peptide Synthesizer (PTI Technologies, Inc.) performs fully automated synthesis of 12 peptides simultaneously with synthesis scales ranging from 0.025 mmol to 0.1 mmol
- The 433A Peptide Synthesizer (Applied Biosystems) is a single column peptide synthesizer that operates in “batch” mode. Due to the on-line conductivity reaction monitoring it is an alternative instrument for fully automated synthesis of peptides with difficult sequences at 0.1-1 mmol scales
- Two preparative HPLC instruments (Beckman-Coulter and Rainin) are used for preparative peptide purification of 15-100 mg peptide per injection
- A Rainin Semipreparative HPLC is used for purifications of smaller quantities (2mg-15 mg of peptide)
- A Beckman Analytical HPLC is used for quality control and for determining optimum specifications for the purification of synthetic peptides
- An Ultimate capillary HPLC equipped for capillary flow (Dionex) is also available
Matrix Assisted Laser Desorption Time Of Flight mass spectrometer with Masslynx version 3.4 software (MALDI-TOF; TOFSpec 2E, Micromass). The three-year-old MALDI TOF is the instrument of choice for analysis of proteins, protein complexes and peptide mixtures generated from protein cleavage (peptide mass mapping). A resolution of 10,000 is typically achieved in reflectron mode for molecular masses below 5000 Da. This configuration is ideal for peptide mass mapping. Linear mode is used for peptides and proteins with molecular masses greater than 5000 Da, and a practical upper limit to the scan range is m/z 200,000.
LCQ Ion Trap mass spectrometer with Excalibur version 1.2 software (Duo, ThermoFinnigan). The 4-year-old ion trap is also equipped with electrospray ionization but provides MS analysis of proteins and peptides at higher resolution than the Platform. It has a scan range of m/z 50-2000. Although this instrument has MS/MS capability, it is primarily needed for quality control of synthetic peptides.
Edman Sequence analysis is performed on a Procise 494HT fully automated instrument (Applied Biosystems).
Circular dichroism analysis is performed on a Jasco J810 state-of-the-art Spectropolarimeter with fluorescence emission detector. This allows for analysis of fluorescence labeled proteins and peptides.
All instruments listed are maintained by trained facility personnel and manufacturer’s service contracts.
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