Dataset Information

Additional precursor purification in isobaric mass tagging experiments by traveling wave ion mobility separation (TWIMS).

ABSTRACT: We investigated if additional peptide purification by traveling wave ion mobility separation (TWIMS) can reduce precursor contamination using a mixture of S. cerevisiae and HeLa proteomes. In accordance with previous reports on FAIMS-Orbitrap instruments, we find that TWIMS provides a remarkable improvement (on average 2.85 times) in signal to noise ratio for sequence ions. We also report, that TWIMS reduces reporter ions contamination by around one third (to 14 - 15% contamination) and even further (to 6-9%) when combined with narrowed quadrupole isolation window.

INSTRUMENT(S): Synapt MS

ORGANISM(S): Homo Sapiens (human) Saccharomyces Cerevisiae (baker's Yeast)

SUBMITTER: Pavel Shliaha

LAB HEAD: Kathryn Susan Lilley

PROVIDER: PXD001047 | Pride | 2014-06-10

REPOSITORIES: Pride

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Dataset's files

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			Action	DRS
	2nd_Manchester_unfractionated_2_5ul_narrow_quad_01.raw.zip	Raw
	2nd_Manchester_unfractionated_2_5ul_narrow_quad_01_stripped.csv	Csv
	2nd_Manchester_unfractionated_2_5ul_narrow_quad_01_stripped.mgf	Mgf
	2nd_Manchester_unfractionated_2_5ul_narrow_quad_02.raw.zip	Raw
	2nd_Manchester_unfractionated_2_5ul_narrow_quad_02_stripped.csv	Csv

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Additional precursor purification in isobaric mass tagging experiments by traveling wave ion mobility separation (TWIMS).

Shliaha Pavel V PV Jukes-Jones Rebekah R Christoforou Andy A Fox Jonathan J Hughes Chris C Langridge James J Cain Kelvin K Lilley Kathryn S KS

Journal of proteome research 20140616 7

Despite the increasing popularity of data-independent acquisition workflows, data-dependent acquisition (DDA) is still the prevalent method of LC-MS-based proteomics. DDA is the basis of isobaric mass tagging technique, a powerful MS2 quantification strategy that allows coanalysis of up to 10 proteomics samples. A well-documented limitation of DDA, however, is precursor coselection, whereby a target peptide is coisolated with other ions for fragmentation. Here, we investigated if additional pept ...[more]

PMID: 24854137

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Project description:The transcriptome, proteome and metabolome of 16 S.cerevisiae backgrounds, combinatorially perturbed in HIS3, LEU2, URA3 and MET15strains are compared. The project was jointly supervised by Kathryn Lilley and Markus Ralser. Transcriptome data has been deposited at ArrayExpress under accession <a href="http://www.ebi.ac.uk/microarray-as/aer/result?queryFor=Experiment&eAccession=E-MTAB-3991">E-MTAB-3991</a>. Metabolome data has been deposited at Metabolights with accession number <a href="http://www.ebi.ac.uk/metabolights/MTBLS168">MTBLS168</a>. The regulation of gene expression in response to nutrient availability is fundamental to the genotype–phenotype relationship. The metabolic–genetic make-up of the cell, as reflected in auxotrophy, is hence likely to be a determinant of gene expression. Here, we address the importance of the metabolic–genetic background by monitoring transcriptome, proteome and metabolome in a repertoire of 16 Saccharomyces cerevisiae laboratory backgrounds, combinatorially perturbed in histidine, leucine, methionine and uracil biosynthesis. The metabolic background affected up to 85% of the coding genome. Suggesting widespread confounding, these transcriptional changes show, on average, 83% overlap between unrelated auxotrophs and 35% with previously published transcriptomes generated for non-metabolic gene knockouts. Background-dependent gene expression correlated with metabolic flux and acted, predominantly through masking or suppression, on 88% of transcriptional interactions epistatically. As a consequence, the deletion of the same metabolic gene in a different background could provoke an entirely different transcriptional response. Propagating to the proteome and scaling up at the metabolome, metabolic background dependencies reveal the prevalence of metabolism-dependent epistasis at all regulatory levels. Urging a fundamental change of the prevailing laboratory practice of using auxotrophs and nutrient supplemented media, these results reveal epistatic intertwining of metabolism with gene expression on the genomic scale.

Dataset Information

Additional precursor purification in isobaric mass tagging experiments by traveling wave ion mobility separation (TWIMS).

Dataset's files

Publications

Additional precursor purification in isobaric mass tagging experiments by traveling wave ion mobility separation (TWIMS).

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