Milk Bottom-Up Proteomics: Method Optimization

Frontiers in Genetics
Delphine VincentSimone Rochfort

Abstract

Milk is a complex fluid whose proteome displays a diverse set of proteins of high abundance such as caseins and medium to low abundance whey proteins such as ß-lactoglobulin, lactoferrin, immunoglobulins, glycoproteins, peptide hormones, and enzymes. A sample preparation method that enables high reproducibility and throughput is key in reliably identifying proteins present or proteins responding to conditions such as a diet, health or genetics. Using skim milk samples from Jersey and Holstein-Friesian cows, we compared three extraction procedures which have not previously been applied to samples of cows' milk. Method A (urea) involved a simple dilution of the milk in a urea-based buffer, method B (TCA/acetone) involved a trichloroacetic acid (TCA)/acetone precipitation, and method C (methanol/chloroform) involved a tri-phasic partition method in chloroform/methanol solution. Protein assays, SDS-PAGE profiling, and trypsin digestion followed by nanoHPLC-electrospray ionization-tandem mass spectrometry (nLC-ESI-MS/MS) analyses were performed to assess their efficiency. Replicates were used at each analytical step (extraction, digestion, injection) to assess reproducibility. Mass spectrometry (MS) data are available via ProteomeXc...Continue Reading

References

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Citations

Mar 15, 2016·Expert Opinion on Drug Discovery·Ivan Fraietta, Fabio Gasparri
Sep 27, 2019·Proteomes·Delphine VincentGerman Spangenberg
Jan 8, 2019·Journal of Dairy Science·Sabrina L Greenwood, Mallory C Honan
Nov 14, 2019·International Journal of Molecular Sciences·Delphine VincentGerman Spangenberg
Jun 11, 2020·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Aliah Zannierah MohsinFarina Mustaffa-Kamal
Feb 20, 2019·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Delphine VincentGerman Spangenberg
Oct 31, 2018·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Delphine VincentSimone Rochfort
Jun 11, 2020·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Muzammeer MansorNuzul Noorahya Jambari
Apr 6, 2021·Comprehensive Reviews in Food Science and Food Safety·Hina KamalAsgar Ali

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Datasets Mentioned

BETA
PXD002529

Methods Mentioned

BETA
Solid Phase Extraction
protein assay
deamination
Assay
PCA
protein extraction

Software Mentioned

Basic Local Alignment Search Tool ( BLAST )
Genedata Expressionist Refiner
Excel
Analyst
Genedata Expressionist Analyst
MASCOT
Proteome Discoverer

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