Proteomic-based analytical approach for the characterization of glutenin subunits in durum wheat.
One of the main objectives of wheat glutenin subunit (GS) analysis is the identification of protein components linked to wheat quality. The proteomic characterization of glutenin has to consider the relatively low levels of arginine and lysine residues and the close sequence similarity among the different groups of these subunits, which hinders or even prevents the identification of the GS. In this study, a proteomic approach has been applied to resolve the heterogeneity of wheat glutenin components. Proteins extracted from Triticum durum flour were first analyzed by two-dimensional gel electrophoresis, which greatly reduced glutenin complexity. The identity of each spot was confirmed by nano liquid chromatography tandem mass spectrometry analysis of tryptic peptides. In parallel, measurements of the high mass range by matrix-assisted laser desorption/ionization time-of-flight analysis allowed detection of the large tryptic peptides. Gathering all data from search engine interrogation, very high sequence coverage was obtained for high molecular weight GS, including Bx7 and By8, in agreement with the known genetic profile of durum wheat. In addition, a truncated form of By8, never detected before, was also found. Low molecular weight GS (LMW-GS) B-type was identified with reasonable sequence coverage, while a clear identification of LWM-GS C- and D-type was hindered by the incompleteness of the wheat DNA databases. This study represents the first comprehensive analysis of the glutenin proteome and provides a reliable method for classifying wheat varieties according to their glutenin profile.
Mamone
Gianfranco
G
Institute of Food Science, CNR, Avellino, Italy. mamone@isa.cnr.it
De Caro
Salvatore
S
Di Luccia
Aldo
A
Addeo
Francesco
F
Ferranti
Pasquale
P
D002853:Chromatography, Liquid / Q000379:methods; D015180:Electrophoresis, Gel, Two-Dimensional; D005433:Flour / Q000032:analysis; D005983:Glutens / Q000032:analysis* / Q000737:chemistry; D008970:Molecular Weight; D021122:Protein Subunits / Q000032:analysis* / Q000737:chemistry; D040901:Proteomics / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods; D014908:Triticum / Q000737:chemistry*; D014357:Trypsin
D021122:Protein Subunits; D005983:Glutens; D014357:Trypsin; C037302:glutenin