Lactoferrin, a protein showing an array of biochemical properties, including immuno-modulation, iron-binding ability, as well as antioxidant, antibacterial and antiviral activities, but which may also represent a potential milk allergen,was isolated from donkey milk by ion exchange chromatography.The characterization of itsprimary structure, by means of enzymatic digestions, SPITC derivatization of tryptic digest,reversed-phase high performance liquid chromatography,electrospray and matrix-assisted laser desorption/ionization mass spectrometry, is reported. Our results allowed the almost complete characterization of donkey lactoferrin sequence, that, at least for the covered sequence, differs from the horsegenomic deduced sequence (UniProtKB Acc. Nr. O77811)byfive point substitutions located at positions 91 (ArgHis), 328 (ThrIle/Leu), 466 (AlaGly), 642 (AsnSer) and 668 (SerAla). Analysis of the glycosylated protein showed that glycans in donkey lactoferrin are linked to the protein backbone via an amide bond to asparagine residues located at the positions 137, 281 and 476.

Sequence characterization and glycosylation sites identification of donkey milk lactoferrin by multiple enzyme digestions and mass spectrometry

Gallina S;CUNSOLO, VINCENZO;SALETTI, Rosaria;MUCCILLI, VERA;Di Francesco A;FOTI, Salvatore;
2016-01-01

Abstract

Lactoferrin, a protein showing an array of biochemical properties, including immuno-modulation, iron-binding ability, as well as antioxidant, antibacterial and antiviral activities, but which may also represent a potential milk allergen,was isolated from donkey milk by ion exchange chromatography.The characterization of itsprimary structure, by means of enzymatic digestions, SPITC derivatization of tryptic digest,reversed-phase high performance liquid chromatography,electrospray and matrix-assisted laser desorption/ionization mass spectrometry, is reported. Our results allowed the almost complete characterization of donkey lactoferrin sequence, that, at least for the covered sequence, differs from the horsegenomic deduced sequence (UniProtKB Acc. Nr. O77811)byfive point substitutions located at positions 91 (ArgHis), 328 (ThrIle/Leu), 466 (AlaGly), 642 (AsnSer) and 668 (SerAla). Analysis of the glycosylated protein showed that glycans in donkey lactoferrin are linked to the protein backbone via an amide bond to asparagine residues located at the positions 137, 281 and 476.
2016
lactoferrin; sequence characterization; glycosylation sites; donkey milk; mass spectrometry
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/20.500.11769/17837
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