Identification of transglutaminase-mediated deamidation sites in a recombinant alpha-gliadin by advanced mass-spectrometric methodologies.
Celiac disease is a permanent immune-mediated food intolerance triggered by ingestion of wheat gliadins in genetically susceptible individuals. It has been reported that tissue transglutaminase plays an important role in the onset of celiac disease by converting specific glutamine residues within gliadin fragments into glutamic acid residues. This process increases binding affinity of gliadin peptides to HLA-DQ2/DQ8 molecules, thus enhancing the immune response. The aim of the present study was to achieve a detailed structural characterization of modifications induced by transglutaminase on gliadin peptides. Therefore, structural analyses were carried out on a recombinant alpha-gliadin and on a panel of 26 synthetic peptides, overlapping the complete protein sequence. Modified glutamine residues were identified by means of advanced mass-spectrometric methodologies on the basis of MALDI-TOF-MS and tandem mass spectrometry. Results led to the identification of 19 of 94 glutamine residues present in the recombinant alpha-gliadin, which were converted into glutamic acid residues by a transglutaminase-mediated reaction. This allowed us to achieve a global view of the modifications induced by the enzyme on this protein. Furthermore, results gathered could likely be utilized as relevant information for a better understanding of processes leading to T-cell recognition of gliadin peptides involved in celiac disease.
Mazzeo
Maria Fiorella
MF
Centro di Spettrometria di Massa Proteomica e Biomolecolare, Istituto di Scienze dell'Alimentazione del CNR, Avellino, Italy.
De Giulio
Beatrice
B
Senger
Stefania
S
Rossi
Mauro
M
Malorni
Antonio
A
Siciliano
Rosa Anna
RA
D000577:Amides / Q000737:chemistry* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D005903:Gliadin / Q000737:chemistry* / Q000378:metabolism; D006168:Guinea Pigs; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins; D011503:Transglutaminases / Q000737:chemistry* / Q000378:metabolism
D000577:Amides; D011994:Recombinant Proteins; D005903:Gliadin; D011503:Transglutaminases