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MS-based characterization of αs2-casein isoforms in donkey's milk

Authors


  • This article is part of the Journal of Mass Spectrometry special issue entitled “2nd MS Food Day” edited by Gianluca Giorgi.

Rosaria Saletti, Department of Chemical Sciences, University of Catania, Viale A. Doria, 6, I-95125 Catania, Italy. E-mail: rsaletti@unict.it

Abstract

The primary structure of four αs2-casein (CN) isoforms, present as minor components in the dephosphorylated CN fraction of a milk sample collected in Eastern Sicily from an individual donkey belonging to the Ragusano breed at middle lactation stage, was determined, using the known donkey's αs2-CN (GenBank Acc. No. CAV00691; Mr 26 028 Da) as reference. Proteins, with experimentally measured Mr of 25 429, 21 939, 25 203 and 21 713 Da, were isolated by the combined use of reversed-phase high-performance liquid chromatography (RP-HPLC) and two-dimensional polyacrylamide gel electrophoresis. The major spot of each gel, corresponding to a single protein, was digested by trypsin, α-chymotrypsin and endoproteinase Glu-C. The resulting peptide mixtures were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and capillary RP-HPLC/nano-electrospray ionization tandem mass spectrometry, and the data obtained were used for the sequence determination. The isoforms are produced from differential splicing events involving exons 4, 5 and 6 and parts of the exon 17. Copyright © 2012 John Wiley & Sons, Ltd.

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