CHARACTERISATION OF SORGHUM KAFIRIN 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 differentMrs, in addition to γ-, α1-, α2- and β-kafirin. These oligomers comprised γ-, α1- and α2-kafirin linked together by disulphide (SS) bonds, as demonstrated by SDS–PAGE analysis after reduction. In contrast, β-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 γ- and α1-kafirin were also detected, whereas monomeric β-kafirin was absent. The polymers comprised γ-, α1-, and β-kafirin, but α2-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 γ- and α1-kafirin of α2- (a «chain terminator») and β-kafirin (a «chain extender»).