Using guanidine-hydrochloride for fast and efficient protein digestion and single-step affinity-purification mass spectrometry

Research output: Contribution to journalJournal articlepeer-review

Standard

Using guanidine-hydrochloride for fast and efficient protein digestion and single-step affinity-purification mass spectrometry. / Poulsen, Jon Wriedt; Madsen, Christian Toft; Young, Clifford; Poulsen, Flemming Martin; Nielsen, Michael L.

In: Journal of Proteome Research, Vol. 12, No. 2, 2013, p. 1020-30.

Research output: Contribution to journalJournal articlepeer-review

Harvard

Poulsen, JW, Madsen, CT, Young, C, Poulsen, FM & Nielsen, ML 2013, 'Using guanidine-hydrochloride for fast and efficient protein digestion and single-step affinity-purification mass spectrometry', Journal of Proteome Research, vol. 12, no. 2, pp. 1020-30. https://doi.org/10.1021/pr300883y

APA

Poulsen, J. W., Madsen, C. T., Young, C., Poulsen, F. M., & Nielsen, M. L. (2013). Using guanidine-hydrochloride for fast and efficient protein digestion and single-step affinity-purification mass spectrometry. Journal of Proteome Research, 12(2), 1020-30. https://doi.org/10.1021/pr300883y

Vancouver

Poulsen JW, Madsen CT, Young C, Poulsen FM, Nielsen ML. Using guanidine-hydrochloride for fast and efficient protein digestion and single-step affinity-purification mass spectrometry. Journal of Proteome Research. 2013;12(2):1020-30. https://doi.org/10.1021/pr300883y

Author

Poulsen, Jon Wriedt ; Madsen, Christian Toft ; Young, Clifford ; Poulsen, Flemming Martin ; Nielsen, Michael L. / Using guanidine-hydrochloride for fast and efficient protein digestion and single-step affinity-purification mass spectrometry. In: Journal of Proteome Research. 2013 ; Vol. 12, No. 2. pp. 1020-30.

Bibtex

@article{9f83ae5809904ca79e9fcfeb69da9c43,
title = "Using guanidine-hydrochloride for fast and efficient protein digestion and single-step affinity-purification mass spectrometry",
abstract = "Protein digestion is an integral part of the {"}shotgun{"} proteomics approach and commonly requires overnight incubation prior to mass spectrometry analysis. Quadruplicate {"}shotgun{"} proteomic analysis of whole yeast lysate demonstrated that Guanidine-Hydrochloride (Gnd-HCl) protein digestion can be optimally completed within 30 min with endoprotease Lys-C. No chemical artifacts were introduced when samples were incubated in Gnd-HCl at 95 °C, making Gnd-HCl an appropriate digestion buffer for shotgun proteomics. Current methodologies for investigating protein-protein interactions (PPIs) often require several preparation steps, which prolongs any parallel operation and high-throughput interaction analysis. Gnd-HCl allow the efficient elution and subsequent fast digestion of PPIs to provide a convenient high-throughput methodology for affinity-purification mass spectrometry (AP-MS) experiments. To validate the Gnd-HCl approach, label-free PPI analysis of several GFP-tagged yeast deubiquitinating enzymes was performed. The identification of known interaction partners demonstrates the utility of the optimized Gnd-HCl protocol that is also scalable to the 96 well-plate format.",
author = "Poulsen, {Jon Wriedt} and Madsen, {Christian Toft} and Clifford Young and Poulsen, {Flemming Martin} and Nielsen, {Michael L}",
year = "2013",
doi = "10.1021/pr300883y",
language = "English",
volume = "12",
pages = "1020--30",
journal = "Journal of Proteome Research",
issn = "1535-3893",
publisher = "American Chemical Society",
number = "2",

}

RIS

TY - JOUR

T1 - Using guanidine-hydrochloride for fast and efficient protein digestion and single-step affinity-purification mass spectrometry

AU - Poulsen, Jon Wriedt

AU - Madsen, Christian Toft

AU - Young, Clifford

AU - Poulsen, Flemming Martin

AU - Nielsen, Michael L

PY - 2013

Y1 - 2013

N2 - Protein digestion is an integral part of the "shotgun" proteomics approach and commonly requires overnight incubation prior to mass spectrometry analysis. Quadruplicate "shotgun" proteomic analysis of whole yeast lysate demonstrated that Guanidine-Hydrochloride (Gnd-HCl) protein digestion can be optimally completed within 30 min with endoprotease Lys-C. No chemical artifacts were introduced when samples were incubated in Gnd-HCl at 95 °C, making Gnd-HCl an appropriate digestion buffer for shotgun proteomics. Current methodologies for investigating protein-protein interactions (PPIs) often require several preparation steps, which prolongs any parallel operation and high-throughput interaction analysis. Gnd-HCl allow the efficient elution and subsequent fast digestion of PPIs to provide a convenient high-throughput methodology for affinity-purification mass spectrometry (AP-MS) experiments. To validate the Gnd-HCl approach, label-free PPI analysis of several GFP-tagged yeast deubiquitinating enzymes was performed. The identification of known interaction partners demonstrates the utility of the optimized Gnd-HCl protocol that is also scalable to the 96 well-plate format.

AB - Protein digestion is an integral part of the "shotgun" proteomics approach and commonly requires overnight incubation prior to mass spectrometry analysis. Quadruplicate "shotgun" proteomic analysis of whole yeast lysate demonstrated that Guanidine-Hydrochloride (Gnd-HCl) protein digestion can be optimally completed within 30 min with endoprotease Lys-C. No chemical artifacts were introduced when samples were incubated in Gnd-HCl at 95 °C, making Gnd-HCl an appropriate digestion buffer for shotgun proteomics. Current methodologies for investigating protein-protein interactions (PPIs) often require several preparation steps, which prolongs any parallel operation and high-throughput interaction analysis. Gnd-HCl allow the efficient elution and subsequent fast digestion of PPIs to provide a convenient high-throughput methodology for affinity-purification mass spectrometry (AP-MS) experiments. To validate the Gnd-HCl approach, label-free PPI analysis of several GFP-tagged yeast deubiquitinating enzymes was performed. The identification of known interaction partners demonstrates the utility of the optimized Gnd-HCl protocol that is also scalable to the 96 well-plate format.

U2 - 10.1021/pr300883y

DO - 10.1021/pr300883y

M3 - Journal article

C2 - 23186134

VL - 12

SP - 1020

EP - 1030

JO - Journal of Proteome Research

JF - Journal of Proteome Research

SN - 1535-3893

IS - 2

ER -

ID: 46282673