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Ac-IP tag enables multiplexed proteome quantification in data-independent acquisition mode

ABSTRACT: We propose the isotopically labeled Ac-IP (acetyl-isoleucine-proline) tag for multiplexed quantification of peptides at the MS1 level in the data-independent acquisition (DIA) mode. Differentially labeled peptides have distinct precursor ions carrying the quantitative information but identical MS2 spectra. The isotopically labeled Ac-Ile part leaves as a neutral loss upon collision-induced dissociation (CID) while fragmentation of the peptide backbone generates information for peptide identification.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Saccharomyces Cerevisiae (baker's Yeast)

SUBMITTER: Xiaobo Tian

LAB HEAD: Rainer Bischoff

PROVIDER: PXD022606 | Pride | 2021-06-07

REPOSITORIES: Pride

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			Action	DRS
	Ac-IP-LysC-yeast-111.mgf	Mgf
	Ac-IP-LysC-yeast-111.mzid.gz	Mzid
	Ac-IP-LysC-yeast-111.raw	Raw
	Ac-IP-LysC-yeast-521-BSA-1051.mgf	Mgf
	Ac-IP-LysC-yeast-521-BSA-1051.mzid.gz	Mzid

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Publications

The Isotopic Ac-IP Tag Enables Multiplexed Proteome Quantification in Data-Independent Acquisition Mode.

Tian Xiaobo X de Vries Marcel P MP Permentier Hjalmar P HP Bischoff Rainer R

Analytical chemistry 20210531 23

Data-independent acquisition (DIA) is an increasingly used approach for quantitative proteomics. However, most current isotope labeling strategies are not suitable for DIA as they lead to more complex MS2 spectra or severe ratio distortion. As a result, DIA suffers from a lower throughput than data-dependent acquisition (DDA) due to a lower level of multiplexing. Herein, we synthesized an isotopically labeled acetyl-isoleucine-proline (Ac-IP) tag for multiplexed quantification in DIA. Differenti ...[more]

PMID: 34053216

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A Collision-induced Dissociation Cleavable Isobaric Tag for Peptide Fragment Ion-based Quantification in Proteomics

Project description:Fragment ion-based proteome quantification methods quantify peptides based on unique peptide fragment ions avoiding the issue of ratio distortion that is commonly observed for reporter ion-based quantification approaches. Herein, we present a novel approach that relies on a collision-induced dissociation (CID)-cleavable isobaric acetyl-isoleucine-proline-glycine (Ac-IPG) tag, which conserves the merits of quantifying peptides based on unique fragment ions while reducing the complexity of the b-ion series thus facilitating data processing. Multiplex labelling is based on selective N-terminal dimethylation followed by derivatizing the C-terminal Lys residue of LysC peptides with isobaric Ac-IPG tags with different isotope distributions on Pro-Gly and Ac-Ile. Upon fragmentation between Ile and Pro, the resulting y-ions with the neutral loss of Ac-Ile can be distinguished between the different labelling channels based on different numbers of isotope labels on the Pro-Gly part. The peak intensities of y-ions contain the information for relative quantification.

2020-08-17 | PXD018790 | Pride

A Versatile Isobaric Tag Enables Proteome Quantification in Data Dependent and Data Independent Acquisition Mode

Project description:Quantifying proteins based on peptide-coupled reporter-ions is a leading multiplexed quantitative strategy in proteomics that significantly alleviates the problem of ratio distortion caused by peptide co-fragmentation, as commonly observed in other reporter-ion based approaches, such as TMT and iTRAQ. Fueled by improvements in mass spectrometry and data processing, data-independent acquisition (DIA) is an attractive alternative to data dependent acquisition (DDA) due to its better reproducibility. While multiplexed labeling is widely used in DDA, it is rarely used in DIA, presumably because current approaches lead to more complex MS2 spectra or to a reduction in quantification accuracy and precision. Herein, we present a versatile acetyl-alanine-glycine (Ac-AG) tag which conceals quantitative information in isobarically labeled peptides and reveals it upon tandem MS in the form of peptide-coupled reporter-ions. Since the peptide-coupled reporter-ion is precursor-specific while fragment ions of the peptide backbone originating from different labeling channels are same, the Ac-AG tag is compatible with both the widely adopted DDA as well as with the DIA mode. By isolating the monoisotopic peak of the precursor ion in DDA, intensities of the peptide-coupled reporter-ions simply represent the relative ratios between constituent samples. While in DIA, the ratio can be inferred after deconvoluting peptide-coupled reporter-ions. The proteome quantification capability of the Ac-AG tag was demonstrated by triplex labeling of a yeast proteome spiked with bovine serum albumin (BSA) over a 10-fold dynamic range. Within a complex proteomics background, the BSA spiked at 1:5:10 ratios was detected at ratios of 1.00 : 4.87 : 10.13 in DDA and 1.16 : 5.20 : 9.64 in DIA.

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2020-04-27 | PXD018589 | Pride

A DNA polymerase ? accessory protein, Mcl1, is required for propagation of centromere structures in fission yeast

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