Journal of Cereal Science vol:60 issue:1 pages:131-137
The high molecular weight glutenin subunits (HMW-GS) play a key role in end-use quality of wheat. Their particular primary structure is mostly derived from DNA sequencing, which gives no information on potential post-translational modifications. This paper reveals the primary structure of HMW-GS 1Dx2 by proteomic analysis. For this purpose, HMW-GS were first isolated from wheat flour (cv. Contra). The relative molecular mass (Mr) of subunit 1Dx2 present in the HMW-GS mixture was then very accurately determined with high-performance liquid chromatography–electrospray ionization-mass spectrometry using a quadrupole-time-of-flight mass analyzer (HPLC–ESI-QTOF-MS). The obtained Mr value (87,105) differed from the value derived from its protein sequence in the NCBI database (87,007). The subunit was further purified by preparative reversed-phase HPLC and partially hydrolyzed with chymotrypsin. The resulting 1Dx2 peptides were then analyzed by HPLC–ESI-MS/MS and the MS data were compared to amino acid sequences in protein databases. The discrepancy between the calculated and the measured Mr of 1Dx2 was explained by a missing proline in the 1Dx2 amino acid sequence from the database and not by any post-translational glycosylation.