A robust protocol for protein extraction and digestion

Michelle Atallah, Mark Flory, Parag Mallick

Research output: Chapter in Book/Report/Conference proceedingChapter

1 Citation (Scopus)

Abstract

Proteins play a key role in all aspects of cellular homeostasis. Proteomics, the large-scale study of proteins, provides in-depth data on protein properties, including abundances and post-translational modification states, and as such provides a rich avenue for the investigation of biological and disease processes. While proteomic tools such as mass spectrometry have enabled exquisitely sensitive sample analysis, sample preparation remains a critical unstandardized variable that can have a significant impact on downstream data readouts. Consistency in sample preparation and handling is therefore paramount in the collection and analysis of proteomic data. Here we describe methods for performing protein extraction from cell culture or tissues, digesting the isolated protein into peptides via in-solution enzymatic digest, and peptide cleanup with final preparations for analysis via liquid chromatography-mass spectrometry. These protocols have been optimized and standardized for maximum consistency and maintenance of sample integrity.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages1-10
Number of pages10
DOIs
StatePublished - Jan 1 2017
Externally publishedYes

Publication series

NameMethods in Molecular Biology
Volume1550
ISSN (Print)1064-3745

Fingerprint

Proteolysis
Proteomics
Proteins
Mass Spectrometry
Biological Phenomena
Peptides
Post Translational Protein Processing
Liquid Chromatography
Homeostasis
Cell Culture Techniques
Maintenance

Keywords

  • Acetone precipitation
  • Enzymatic solution digest
  • Liquid chromatography-mass spectrometry
  • Protein extraction
  • Proteomics

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

Cite this

Atallah, M., Flory, M., & Mallick, P. (2017). A robust protocol for protein extraction and digestion. In Methods in Molecular Biology (pp. 1-10). (Methods in Molecular Biology; Vol. 1550). Humana Press Inc.. https://doi.org/10.1007/978-1-4939-6747-6_1

A robust protocol for protein extraction and digestion. / Atallah, Michelle; Flory, Mark; Mallick, Parag.

Methods in Molecular Biology. Humana Press Inc., 2017. p. 1-10 (Methods in Molecular Biology; Vol. 1550).

Research output: Chapter in Book/Report/Conference proceedingChapter

Atallah, M, Flory, M & Mallick, P 2017, A robust protocol for protein extraction and digestion. in Methods in Molecular Biology. Methods in Molecular Biology, vol. 1550, Humana Press Inc., pp. 1-10. https://doi.org/10.1007/978-1-4939-6747-6_1
Atallah M, Flory M, Mallick P. A robust protocol for protein extraction and digestion. In Methods in Molecular Biology. Humana Press Inc. 2017. p. 1-10. (Methods in Molecular Biology). https://doi.org/10.1007/978-1-4939-6747-6_1
Atallah, Michelle ; Flory, Mark ; Mallick, Parag. / A robust protocol for protein extraction and digestion. Methods in Molecular Biology. Humana Press Inc., 2017. pp. 1-10 (Methods in Molecular Biology).
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