UC Davis Proteomics Facility - Protocols, How To's, sample Data and Presentations

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1D, 2D and 3D Peptide LC MS/MS Analysis of Low Abundance Proteins in Complex Proteomes

Submitter: brettsp	Released: Thu, 12-Jun-2008

Download Now Due to the large number of proteins and wide range of relative abundances in complex proteome samples, a significant amount of resolution is required prior to MS analysis in order to identify, characterize and/or quantitate low abundance proteins. Proteomics researchers currently use a wide variety of separation protocols to separate proteins and/or peptides prior to MS analysis (1D or 2D gels, 1D or 2D LC) but compromises are made to minimize losses and total analysis time. This study introduces a new automated 3D peptide separation protocol that offers maximum peptide resolution and recovery for a given analysis time. D1 utilizes high pH RPLC to desalt (necessary prior to SCX) and fractionate the digest. Fractions from D1 (pH adjusted to 3) are further fractionated by D2 SCX (which removes detergents and excess acetonitrile) prior to desalting and D3 low pH RP LC-MS/MS. Using a new ESI source that provides high sensitivity over a wide flow range, we were able to do an optimized comparison of 1D, 2D and 3D protocols to determine the relative advantages and disadvantages of these different separation schemes within a fixed MS/MS acquisition time.	Version: 0 Downloads152 File Size: 0 bytes Platform: None Home Page: Not Specified Rating: (0 Votes)

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ABRF 2008 Facility Poster

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Submitter: brettsp	Released: Thu, 12-Jun-2008

Download Now acid analysis, Edman sequencing and tandem mass spectrometry techniques including protein identification, post-translational modification discovery, label free quantitation and protein cross-linking analysis. The facility utilizes state of the art instrumentation, including a Thermo LTQ-FT Ultra. Our group utilizes open source software and uses techniques and protocols that are made publicly available. Using open source software allows users to analyze their own data without significant monetary investment. We also offer data analysis and sample preparation classes to UC Davis and the community at large, including a week long hands on summer short course. Our group operates as an open core facility withithe Genome Center on the UC Davis campus	Version: 0 Downloads87 File Size: 0 bytes Platform: None Home Page: Not Specified Rating: (0 Votes)

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Agilent LC-CHIP Trap and Orbi-Trap Data set

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Submitter: brettsp	Released: Wed, 28-Sep-2005

Download Now Same sample run on Agilents new LC-Chip Ultra trap and a Orbi-trap. Not sure if you can read too much into this comparison as the gradients and run conditions were a bit different, althought not much.	Version: 0 Downloads377 File Size: 84.49 KB Platform: None Home Page: Not Specified Rating: (0 Votes)

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Automating x!tandem and Scaffold on a Large Scale Computing Cluster

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Submitter: brettsp	Released: Thu, 12-Jun-2008

Download Now Many scientific centers and life science facilities today have access to medium to large computer clusters or large numbers of computers that can be employed in a grid type computing environment. Often these clusters or grids employ some sort of batch queuing and job management system. X!tandem , an open source protein identification program has been adapted to run on these environments by using Sun Grid Engine (SGE) as its job management system for submitting X!tandem search jobs. Both X!tandem and SGE are open source platforms allowing life scientists to fully utilize their computing resources without the significant investment typically required by proprietary solutions. Here we describe an easy to use web interface that allows X!tandem jobs to be submitted to computer clusters for scalable computing. The results can be retrieved without any complicated interactions from the end user. The web interface was modified based on thegpm project (www.thegpm.org). In addition we have written the interface and backend applications to automatically create scaffold (www.proteomesftware.com) files from the X!tandem search output using the commercial program scaffold batch. All of the software written for this project will be made available under an open source license.	Version: 0 Downloads91 File Size: 0 bytes Platform: None Home Page: Not Specified Rating: (0 Votes)

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AutoMSN3

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Submitter: brettsp	Released: Mon, 06-Feb-2006

Download Now Small perscript that calls extract_msn to process a whole directory of raw files and turns them into MGF files. It deletes all of the dta and out files (you never see them, yea!!!)	Version: 0 Downloads243 File Size: 9.23 KB Platform: Windows Home Page: Not Specified Rating: (0 Votes)

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c1 Fragment Ions in Collision-induced Dissociation of Glutamine Containing

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Submitter: brettsp	Released: Thu, 12-Jun-2008

Download Now Unusually stable c1 fragment ion was observed when glutamine is at the second position from the N-terminus c1 fragment is generated independent of the kind of the N-terminal residue of the peptide, but the intensity was strongly dependent on its side chain. This information could be used to sequence the first two amino acid residues from N-terminus and consequently to improve protein identification in de novo sequencing-blast/fasta search. A consecutive fragmentation mechanism is proposed with six-membered ring b2 ion as an intermediate to explain this exceptional stability of c1 fragment ion. Introduction In the interpretation of CID spectrum, b and y series ions are most commonly used to deduce the sequence information. Accordingly, fragment ions other than b and y series are often not fully accounted when their mass differences are not matched with any amino acid residue mass. We present here a fragment ion of a non-b, non-y that might be useful in de novo sequencing of peptides. c1 ion was observed with significant yield in the MS/MS spectra of peptide ions containing glutamine as the second amino acid residue from the N-terminus. The utility of this information	Version: 0 Downloads77 File Size: 0 bytes Platform: None Home Page: Not Specified Rating: (0 Votes)

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Cross-linking Site Mapping of Sindbis Virus Structural Proteins

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Submitter: brettsp	Released: Thu, 12-Jun-2008

Download Now Overview High-throughput approach (nanoLC-MS/MS and “X-link” search) was used for the cross-linking site mapping of Sindbis virus structural proteins. Computer program (“X-link”) was developed to find matching sequences from thousands of MS/MS spectra. Identified E1/E2, E1/E1, and E2/E2 cross-linked peptides using BS3 and DSS cross-linkers. Also found four intra-peptide links. Introduction Sindbis virus (SV) is an alphavirus, which is propagated in nature via a complicated life cycle involving insect vectors and mammalian hosts. The outer protein shell is composed of the E1 and E2 glycoproteins organized in trimers of heterodimers. However, it is not well known how these structural proteins interact with each other to form outer shell. In this study, we use cross linking agents to covalently stabilize the interacting parts of the proteins and attempt to identify the cross linking sites. This study allows us to locate areas where these proteins interact with each other or with another copy of themselves and allow us to gain both structural and possibly functional information about the virus.	Version: 0 Downloads86 File Size: 0 bytes Platform: None Home Page: Not Specified Rating: (0 Votes)

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Cryptococcus neoformans: Faster Methods to Characterize the Extracellular Fungal Proteome

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Submitter: brettsp	Released: Thu, 12-Jun-2008

Download Now To further elucidate the extracellular proteome of C. neoformans, we collected both secreted and proteolytically-released proteins from fungal cells in vivo, removed small molecules and lipids, then reduced and alkylated for CLC-MS/MS analysis. We also broke C. neoformans cells mechanically and compared the sequence coverage to the secreted and enzymatically-released proteins.	Version: 0 Downloads85 File Size: 0 bytes Platform: None Home Page: Not Specified Rating: (0 Votes)

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current citations

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Submitter: brettsp	Released: Tue, 30-May-2006

Download Now current citations	Version: 0 Downloads295 File Size: 8.35 KB Platform: None Home Page: Not Specified Rating: (0 Votes)

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E-Values

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Submitter: brettsp	Released: Tue, 07-Feb-2006

Download Now A short slide about what e-Values are. Not sure who I stole this from…	Version: 0 Downloads258 File Size: 11.21 KB Platform: None Home Page: Not Specified Rating: (0 Votes)

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