isoScale

isoScale quantifies co-fragmenting isobaric peptides in MS/MS spectra by leveraging total ion intensity measurements to provide absolute and relative quantification of peptides that differ by post-translational modification localization.

Key Features:

• Co-fragmenting isobaric peptide quantification: Quantifies peptides that co-fragment in MS/MS spectra and differ only by localizations of post-translational modifications (PTMs).
• Total ion intensity-based measurements: Uses total ion intensity measurements from MS/MS spectra for quantification.
• Absolute and relative quantification: Reports both absolute and relative abundances of isobaric peptide species.
• Middle-down proteomics compatibility: Handles large polypeptides in the 5–7 kDa range for sensitive mapping and quantification of coexisting PTMs.
• Chromatography and MS integration: Integrates with a platform using a reverse-phase (RP) trap column followed by weak cation exchange-hydrophilic interaction liquid chromatography interfaced to ESI MS/MS.
• High mass accuracy ETD MS/MS support: Operates with high mass accuracy electron transfer dissociation (ETD) MS/MS sequencing.
• Hypermodified histone N-terminal tail sequencing: Enables separation and sequencing of hypermodified histone N-terminal tails for unambiguous localization of combinatorial PTMs.
• Combinatorial histone mark quantification: Has been used to quantify 256 distinct combinatorial histone marks across histones H3, H4, and H2A and to identify 713 combinatorial marks in purified histone H3 samples.

Scientific Applications:

• Histone modification mapping: Quantifies and maps combinatorial PTMs on histone N-terminal tails in middle-down proteomics datasets.
• Comparative PTM analysis in genetic knockouts: Analyzes PTM abundance changes in murine embryonic stem cells with genetic knockouts, including Suz12(-/-) samples.
• Detection of specific PTM changes: Detected a seven-fold reduction in H3K27me2 and H3K27me3 levels in Suz12(-/-) cells and alterations in the relative abundance of 16 other single PTMs on histone H3.
• Cataloguing combinatorial marks: Enables identification and quantification of hundreds of distinct combinatorial histone marks across H3, H4, and H2A.

Methodology:

Computes absolute and relative abundances of co-fragmenting isobaric peptides from MS/MS spectra using total ion intensity measurements.