Characterisation of sorghum kafirins in relation to their cross-linking behaviour

Sorghum storage proteins (kafirins) were extracted with 60% 2-methyl-propan-2-ol (t-butanol) and analysed in the unreduced form by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The pattern obtained showed the presence of various protein oligomers of different M(r)s, in addition to gamma-, alpha(1)-, alpha(2)- and beta-kafirin. These oligomers comprised gamma-, alpha(1)- and alpha(2)-kafirin linked together by disulphide (SS) bonds, as demonstrated by SDS-PAGE analysis after reduction. In contrast, beta-kafirin was present only in its monomeric form and not as a component of the oligomers. Size-exclusion high-performance liquid chromatography (SE-HPLC) analysis confirmed these results, and indicated that about 70% of the total protein in the extract was in the form of SS-linked oligomers. Sonication of the residue allowed the extraction of additional protein, which, when analysed by SDS-PAGE (in both the unreduced and reduced forms) and SE-HPLC, was shown to comprise mainly polymers of high molecular size, representing 90% of the total protein extracted by sonication. In this case, some monomers and dimers of gamma- and alpha(1)-kafirin were also detected, whereas monomeric beta-kafirin was absent. The polymers comprised gamma-, alpha(1)-, and beta-kafirin, but alpha(2)-kafirin was not detected. From these results, we suggest that the degree of polymerisation of kafirin proteins is determined by the competitive linkage through SS bonds to gamma- and alpha(1)-kafirin of alpha(2)- (a 'chain terminator') and beta-kafirin (a 'chain extender'). (C) 1998 Academic Press.