Dataset Information

Ion mobility tandem mass spectrometry enhances performance of bottom-up proteomics

ABSTRACT: We describe a novel method utilizing ion mobility-based concentration of peptide fragment ions on a qTOF mass spectrometer improving the sensitivity of bottom-up proteomics by up to 10-fold. This enabled the identification of 7,500 human proteins within one day and 8,600 phosphorylation sites within 5h of LC-MS/MS time. The method also proved powerful for multiplexed quantification experiments exemplified by the chemoproteomic interaction analysis of HDACs with Trichostatin A.

INSTRUMENT(S): Synapt MS

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Spleen, Testis, Ovary, Prostate Gland, Lung, Liver, Kidney

SUBMITTER: Hannes Hahne

LAB HEAD: Bernhard Kuster

PROVIDER: PXD000863 | Pride | 2015-08-18

REPOSITORIES: Pride

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Publications

Ion mobility tandem mass spectrometry enhances performance of bottom-up proteomics.

Helm Dominic D Vissers Johannes P C JP Hughes Christopher J CJ Hahne Hannes H Ruprecht Benjamin B Pachl Fiona F Grzyb Arkadiusz A Richardson Keith K Wildgoose Jason J Maier Stefan K SK Marx Harald H Wilhelm Mathias M Becher Isabelle I Lemeer Simone S Bantscheff Marcus M Langridge James I JI Kuster Bernhard B

Molecular & cellular proteomics : MCP 20140808 12

One of the limiting factors in determining the sensitivity of tandem mass spectrometry using hybrid quadrupole orthogonal acceleration time-of-flight instruments is the duty cycle of the orthogonal ion injection system. As a consequence, only a fraction of the generated fragment ion beam is collected by the time-of-flight analyzer. Here we describe a method utilizing postfragmentation ion mobility spectrometry of peptide fragment ions in conjunction with mobility time synchronized orthogonal ion ...[more]

PMID: 25106551

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Project description:Paracoccidioidomycosis (PCM) is an endemic mycosis in Latin America. The PCM is a systemic disease caused by the thermo dimorphic fungus Paracoccidioides spp, which can exist as mycelia and yeast forms. The mycelia form can be found in environment at temperatures between 18 °C and 25 °C and produces spores or conidia. The PCM infection route occurs by the inhalation of conidia. Once in the lungs, these particles are converted into yeasts pathogenic form. Until now, metabolic aspects and virulence factors related to Paracoccidioides lutzii infective propagule are little understood. In this concern, a global proteomic study of P. lutzii conidia was performed. For conidia production, the mycelia of the isolate Pb01 were cultured in potato agar medium during 90 days at 18 °C. Obtained conidia were collected and purified. The proteins were extracted and subjected to tryptic digestion followed by identification by NanoUPLC-MSE approach. We identified a total of 242 proteins in P. lutzii conidia which were subjected to in silico functional analysis Proteins putatively acting as adhesins, such as GAPDH, enolase, and the glycoprotein β-1,3-glucanosyltransferase (gel2) were identified during our analysis. We also identified proteins related to signal transduction pathways, such as Ras and RhoA GTPases, previously demonstrated to be required for morphologic changes in pathogenic fungi. Proteins related to evasion, defense and virulence were also identified. Proteins acting during temperature shifts or oxidative stress provided by the host environment such as HSP90, catalase, mitochondrial peroxiredoxin Prx1 were identified. It were also identified proteins related to energy production and protein biosynthesis, which can provide important aspects to survival of this resting cells. These results highlight that P. lutzii conidia contain proteins that can contribute to its maintenance in the environment and also molecules related to host-pathogen interactions.

Dataset Information

Ion mobility tandem mass spectrometry enhances performance of bottom-up proteomics

Publications

Ion mobility tandem mass spectrometry enhances performance of bottom-up proteomics.

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