Biochemical characterization and quantification of the storage protein (secalin) types in rye flour

Kernels of the rye cultivars Danko and Halo were milled into white flour and compared with flour of the wheat cultivar Rektor. Flour proteins were extracted stepwise with a salt solution (albumins-globulins), 60% ethanol (prolamins), and 50% 2-propanol under reducing conditions (glutelins). The quantification by reversed-phase HPLC indicated that the extractable proteins of both rye flours consisted of approximate to26% albumins-globulins, 65% prolamins, and 9% glutelins. Compared with wheat flour, rye flours comprised significantly higher proportions of nonstorage proteins (albumins-globulins) and lower proportions of polymerized storage proteins (glutelins). SDS-PAGE revealed that the prolamin fractions of rye contained all four storage protein types (HMW, gamma-75k, omega, and gamma-40k secalins), whereas the glutelin fractions contained only HMW and gamma-75k secalins. The quantification of secalin types by RP-HPLC showed a close relationship between the two cultivars.The gamma-75k secalins contributed nearly half (approximate to46%) of the total storage proteins, followed by gamma-40k secalins (24%) and omega secalins (17%); HMW secalins (approximate to7%) were minor components, and 6% of eluted proteins were not identified. The amino acid composition of gamma-40k secalins corresponded to those of gamma-gliadins of wheat, whereas gamma-75k secalins were characterized by higher contents of glutamine and proline. Matrix-assisted laser desorption/ionization and time of flight mass spectrometry (MALDI-TOF MS) indicated molecular masses of about 52,000 (gamma-75k) and 32,000 (gamma-40k), respectively. N-terminal amino acid sequences were homologous with those of wheat gamma-gliadins except for position 5 (asparagine in gamma-75k and glutamine in gamma-40k secalins) and position 12 (cysteine in gamma-75k secalins). The N-terminal amino acid sequences of HMW and omega-secalins were homologous with those of the corresponding protein types of wheat. Gel-permeation HPLC of prolamin fractions revealed that rye flours contained a significantly higher proportion of ethanol-soluble oligomeric proteins than wheat flour.