PROTEOMICS SHARED RESOURCE Timothy Haystead, Ph.D., Director The Proteomics Shared Resource provides state-of-the-art instrumentation and methods to members of the Programs of the Duke Comprehensive Cancer Center. The Facility houses two state of the art mass spectrometers, two automated Edman sequenators, two HPLCs, two FPLCs and an automated tryptic digestion peptide extraction workstation. The primary use of the mass spectrometers is in the identification of proteins and peptides. The ABI MALDI-TOF-TOF instrument enables high throughput protein and peptide identification at a very sensitive level (~fmol). Identifications of analyzed proteins arc carried out by peptide mass finger printing and MS/MS searches. The Facility also houses an ABI QSTAR nano spray mass spectrometer for high sensitivity de novo sequencing of peptides, With the development of the FASTS and TFASTS algorithms by the Haystead laboratory we also offer the unique capability of multipeptide de novo sequence searches of the databases. This service is particularly useful for cross-species analysis with short peptide sequences and searching of poorly annotated DNA databases. Both mass spectrometers can be used to identify post-translational modifications on proteins and peptides, in particular phosphorylation sites using neutral loss (MALDI) or precursor ion scanning (electrospray) methods. Conventional Edman based N terminal sequencing is also available in the Facility for the characterization of recombinant proteins and phosphorylation sites which have been labeled with 32P-orthophosphate. The Facility also provides front-end support both in the form of protein and peptide purification instrumentation (the HPLCs and FPLCs) as well as consultation services. Consults usually revolve around strategies for protein isolation and compatibility of experimental procedures with analysis of proteins and peptides by mass spectrometry.
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