Proteolytic activity in sorghum flour and its interference in protein analysis

An alkaline solvent containing SDS and reducing agent is commonly used for extraction of sorghum and maize proteins before further analysis (SDS-PAGE or enzyme-linked immunosorbant assay [ELISA]) (Wallace et al 1990; Hamaker et al 1995; Oria et al 1995). It is also used as the final extraction solvent in the Landry-Moureaux (1970) protein fractionation method. The solvent composition used by Wallace et al (1990) for extraction of essentially the total grain protein contains 1% SDS and 2% 2-mercaptoethanol (2-ME) in a berate buffer at pH 10. During routine analysis of extracted sorghum proteins using SDS-PAGE, we noticed that, for a few sorghum cultivars, proteins bands appeared hydrolyzed to a significant extent. Bands representing both kafirin (the sorghum storage protein), and nonkafirin proteins were nearly eliminated from the gel, and there was an accumulation of low molecular weight peptides. As this suggested the presence of a protease active under the extraction conditions, we further investigated this possibility. In this brief report, a solvent-active protease is revealed to be present to some degree in most stored sorghum flours. Its origin appears to be fungal. The significance of this finding rests in the interference that this protease causes in sorghum, as well as millet, grain protein analyses.