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Optimized NHS-amine labeling for multiplexed phosphoproteome study.

ABSTRACT: In this project, we report the detrimental effect of hydroxylamine, a quenching reagent for NHS-amine reaction, on phosphopeptides. The degree of phosphopeptide enrichment was greatly improved by a desalting method compared with a vacuum-drying method right after quenching reaction. We demonstrate that vacuum-drying in presence of hydroxylamine promotes β-elimination of phosphate groups from phosphoserine and phosphothreonine.

INSTRUMENT(S): LTQ Orbitrap, Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

DISEASE(S): Brain Cancer

SUBMITTER: Yumi Kwon

LAB HEAD: Cheolju Lee

PROVIDER: PXD008082 | Pride | 2018-02-27

REPOSITORIES: Pride

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			Action	DRS
	160719_TMT_Phospho_Opt1_Exp01.raw	Raw
	160719_TMT_Phospho_Opt1_Exp02.raw	Raw
	160719_TMT_Phospho_Opt1_Exp03.raw	Raw
	160719_TMT_Phospho_Opt1_Exp04.raw	Raw
	160719_TMT_Phospho_Opt1_Exp05.raw	Raw

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Publications

Neutralizing the Detrimental Effect of an N-Hydroxysuccinimide Quenching Reagent on Phosphopeptide in Quantitative Proteomics.

Kwon Yumi Y Ju Shinyeong S Kaushal Prashant P Lee Jin-Won JW Lee Cheolju C

Analytical chemistry 20180214 5

One of the most common chemistries used to label primary amines utilizes N-hydroxysuccinimide (NHS), which is also structurally incorporated in various quantitative proteomic reagents such as isobaric tags for relative and absolute quantification (iTRAQ) and tandem mass tags (TMT). In this paper we report detrimental effect of hydroxylamine, a widely used quenching reagent for excess NHS, on phosphopeptides. We found an impairment in the degree of phosphopeptide identification when hydroxylamine ...[more]

PMID: 29406695

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