Protocol for absolute quantification of proteins in Gram-negative bacteria based on QconCAT-based labeled peptides

Nicolás Gurdo; Shannara Kayleigh Taylor Parkins; Martina Fricano; Tune Wulff; Lars Keld Nielsen; Pablo Iván Nikel

doi:10.1016/j.xpro.2023.102060

Protocol for absolute quantification of proteins in Gram-negative bacteria based on QconCAT-based labeled peptides

Nicolás Gurdo, Shannara Kayleigh Taylor Parkins, Martina Fricano, Tune Wulff, Lars Keld Nielsen, Pablo Iván Nikel^*

^*Corresponding author for this work

Research output: Contribution to journal › Journal article › Research › peer-review

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Abstract

Mass-spectrometry-based absolute protein quantification uses labeled quantification concatamer (QconCAT) as internal standards (ISs). To calculate the amount of protein(s), the ion intensity ratio between the analyte and its cognate IS is compared in each biological sample. The present protocol describes a systematic workflow to design, produce, and purify QconCATs and to quantify soluble proteins in Pseudomonas putida KT2440. Our methodology enables the quantification of detectable peptide and serves as a versatile platform to produce ISs for different biological systems.

Original language	English
Article number	102060
Journal	STAR Protocols
Volume	4
Issue number	1
ISSN	2666-1667
DOIs	https://doi.org/10.1016/j.xpro.2023.102060
Publication status	Published - 2023

Keywords

Biotechnology and Bioengineering
Mass Spectrometry
Microbiology
Protein Biochemistry
Protein Expression and Purification
Systems Biology

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10.1016/j.xpro.2023.102060

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title = "Protocol for absolute quantification of proteins in Gram-negative bacteria based on QconCAT-based labeled peptides",

abstract = "Mass-spectrometry-based absolute protein quantification uses labeled quantification concatamer (QconCAT) as internal standards (ISs). To calculate the amount of protein(s), the ion intensity ratio between the analyte and its cognate IS is compared in each biological sample. The present protocol describes a systematic workflow to design, produce, and purify QconCATs and to quantify soluble proteins in Pseudomonas putida KT2440. Our methodology enables the quantification of detectable peptide and serves as a versatile platform to produce ISs for different biological systems.",

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author = "Nicol{\'a}s Gurdo and {Taylor Parkins}, {Shannara Kayleigh} and Martina Fricano and Tune Wulff and Nielsen, {Lars Keld} and Nikel, {Pablo Iv{\'a}n}",

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year = "2023",

doi = "10.1016/j.xpro.2023.102060",

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AU - Gurdo, Nicolás

AU - Taylor Parkins, Shannara Kayleigh

AU - Fricano, Martina

AU - Wulff, Tune

AU - Nielsen, Lars Keld

AU - Nikel, Pablo Iván

PY - 2023

Y1 - 2023

N2 - Mass-spectrometry-based absolute protein quantification uses labeled quantification concatamer (QconCAT) as internal standards (ISs). To calculate the amount of protein(s), the ion intensity ratio between the analyte and its cognate IS is compared in each biological sample. The present protocol describes a systematic workflow to design, produce, and purify QconCATs and to quantify soluble proteins in Pseudomonas putida KT2440. Our methodology enables the quantification of detectable peptide and serves as a versatile platform to produce ISs for different biological systems.

AB - Mass-spectrometry-based absolute protein quantification uses labeled quantification concatamer (QconCAT) as internal standards (ISs). To calculate the amount of protein(s), the ion intensity ratio between the analyte and its cognate IS is compared in each biological sample. The present protocol describes a systematic workflow to design, produce, and purify QconCATs and to quantify soluble proteins in Pseudomonas putida KT2440. Our methodology enables the quantification of detectable peptide and serves as a versatile platform to produce ISs for different biological systems.

KW - Biotechnology and Bioengineering

KW - Mass Spectrometry

KW - Microbiology

KW - Protein Biochemistry

KW - Protein Expression and Purification

KW - Systems Biology

U2 - 10.1016/j.xpro.2023.102060

DO - 10.1016/j.xpro.2023.102060

M3 - Journal article

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AN - SCOPUS:85146991401

SN - 2666-1667

VL - 4

JO - STAR Protocols

JF - STAR Protocols

IS - 1

M1 - 102060

ER -

Protocol for absolute quantification of proteins in Gram-negative bacteria based on QconCAT-based labeled peptides

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