Dataset Information

A combined gas phase separation strategy for ADP-ribosylated peptides

ABSTRACT: ADP-ribosylation (ADPr) is a posttranslational modification that is best studied using mass spectrometry. Method developments that are permissive to low inputs or baseline levels of protein ribosylation represent the next frontier in the field. High-field asymmetric waveform ion mobility spectrometry (FAIMS) reduces peptide complexity in gas phase, providing a means to achieve maximal ADPr peptide sequencing depth. We therefore investigated the extent to which FAIMS with or without traditional gas-phase fractionation/separation (GPS) can increase the number of ADPr peptides. We examined ADPr peptides enriched from mouse spleens. We gleaned additional insight by also reporting findings from the corresponding non-ADPr peptide contaminants, and the peptide inputs for ADPr peptide enrichment. At increasingly higher compensation voltages, ADPr peptides were more stable whereas the non-ADPr peptides were filtered out. A combination of 3 GPS survey scans, each with 8 compensation voltages resulted in 790 high-confidence ADPr peptides, compared to 90 with GPS alone. A simplified acquisition strategy requiring only two injections corresponding to two MS1 scan ranges coupled to optimized compensation voltage settings, provided 402 ADPr peptides corresponding to 234 ADPr proteins. We conclude that our combined gas phase separation strategy is a valuable addition to any ADP-ribosylome workflow.

INSTRUMENT(S): Orbitrap Exploris 480

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Spleen, Liver

SUBMITTER: Sasha Singh

LAB HEAD: Masanori Aikawa

PROVIDER: PXD040898 | Pride | 2023-08-30

REPOSITORIES: Pride

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Publications

A Combined Gas-Phase Separation Strategy for ADP-ribosylated Peptides.

Kasai Taku T Kuraoka Shiori S Higashi Hideyuki H Delanghe Bernard B Aikawa Masanori M Singh Sasha A SA

Journal of the American Society for Mass Spectrometry 20230817 10

ADP-ribosylation (ADPr) is a post-translational modification that is best studied using mass spectrometry. Method developments that are permissive with low inputs or baseline levels of protein ribosylation represent the next frontier in the field. High-field asymmetric waveform ion mobility spectrometry (FAIMS) reduces peptide complexity in the gas phase, providing a means to achieve maximal ADPr peptide sequencing depth. We therefore investigated the extent to which FAIMS with or without tradit ...[more]

PMID: 37589412

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Project description:Masculinized female Eastern Mosquitofish (Gambusia holbrooki) have resided downstream of paper mills in Florida since the 1980's. The potential impacts of this effluent on the mosquitofish endocrine system are unknown. The objective of this study was to evaluate gene expression patterns of endocrine system genes and global gene expression patterns in female G. holbrooki from a paper mill-impacted site. Masculinized female G. holbrooki were collected from a paper mill-impacted site (Fenholloway River) and from a reference site (Econfina River) and microarray analysis in livers was conducted. Hepatic microarray analysis revealed an increase in the expression of metabolic genes at the Fenholloway, with similarities in individual genes and biological processes compared to G. holbrooki exposed to androgens. These data indicate G. holbrooki from the Fenholloway may be impacted by a mixture of endocrine-active chemicals, including androgens. During the summer of 2012, G. holbrooki were captured from one site downstream of the Buckeye Pulp and Paper Mill (Taylor County, Perry, FL, USA) on the Fenholloway River (GPS coordinates: N 30 058.341’, W 83 588.569’) and one site in the Econfina conservation area (GPS coordinates: N 30 08.549', W 83 51.962') . Only sexually mature G. holbrooki (females > 15cm standard length and with the presence of the gravid spot near the vent) were collected. A 1/8 mesh seine was used for sample collection. Female G. holbrooki were transferred to 5 gallon aerated buckets filled with site water and were processed at the site immediately after collection. Fish were anesthetized using Tricaine-S (Western Chemical, Ferndale, USA) and sacrificed via spinal transection. Oocyte development was assessed upon dissection and livers were removed and stored in RNAlater (Qiagen, Hilden, Germany) overnight at 4 C before storage at -80 C. RNA was isolated from the livers using TRIzol (Invitrogen, Grand Island, USA), hydrated using RNAsecure (Ambion, Grand Island, USA), and DNase treated using the Turbo DNA-free kit (Ambion, Grand Island, USA). Four oocyte-development stage-matched RNA samples per treatment were evaluated for RNA integrity using the 2100 BioAnalyzer (Agilent, Santa Clara, USA). The range of RIN values was 7.8-8.9.

Dataset Information

A combined gas phase separation strategy for ADP-ribosylated peptides

Publications

A Combined Gas-Phase Separation Strategy for ADP-ribosylated Peptides.

Similar Datasets

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