Verification of the cDNA deduced sequence of glutenin subunit 1Dx5 and an Mr 58000 repetitive peptide by matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-MS)

MALDI-MS was used to confirm the cDNA derived amino acid sequence of HMW subunit 1Dx5and an E. coli-expressed repetitive Mr 58 000 peptide based on residues 102–643. Analysis of the Mr58 000 peptide and of tryptic peptides separated by RP-HPLC showed that the experimentallydetermined Mr are consistent, with the calculated values, within the experimental error. This indicatesa substantial correctness of the gene derived sequence. In the same way, analysis of tryptic peptidesfrom 1Dx5 allowed about 80% of the sequence to be confirmed, covering residues 1–669 with theexception of two short peptides. There was also good agreement with sequence of the Mr 58 000peptide in the region of identity. The results, giving evidence for a substantial correctness of a largepart of the sequence, confirm the absence of extensive glycosylation in subunit 1Dx5 and demonstratethe value of MALDI-MS for direct sequence analysis of HMW subunits and similar proteins.