Effects on gluten of heating at different moisture contents. II. Changes in physicochemical properties and secondary structure

When gluten is heated at 80 degrees C for 30 min, dramatic changes are observed in its physico-chemical properties if moisture content exceeds 20%. To understand these changes further we characterised the extractability, sulphydryl/disulphide content, conformation and hydrophobicity of the proteins extensively. Gel-filtration chromatography (GFC) on Sepharose Cl-6B column of a 1.5% (w/v) sodium dodecyl sulphate (SDS) extract of heated gluten resulted in a void volume peak consisting of glutenin aggregates with a relative molecular mass (Mr) greater than 1 X 10(6). This peak was separated further on a Biogel A-150m column into three fractions. The amount of the glutenin fraction with the greatest Mr (Mr > 1 X 10(8)) extracted from fully-denatured gluten was only 5% of that from the unheated sample. Meanwhile, the amounts of free sulphydryl groups were decreased, indicating an increase in disulphide bonds. Labelling studies with a fluorescent label specific for sulphydryl groups in large subunits (Mr > 4 X 10(4) were decreased, but not in the subunits around Mr 2 X 10(1). Heating gluten at a moisture content above 20% also affected the conformation of the glutenin, but not that of the gliadin, albumins and globulins isolated by GFC, as indicated by circular dichroism measurements of proteins extracted in 1.5% (w/v) SDS. The association constant and the number of binding sites of the hydrophobic ligand 8-anilino-naphthalene sulphonic acid decreased when gluten with a moisture content of 13% or higher was heated. The decrease in hydrophobicity may be explained by hypothesizing that, upon heating, hydrophobic groups are exposed to such an extent that the protein aggregates. After aggregation the hydrophobicity is decreased. The results indicate that, after heating gluten at different moisture contents, the first major change to be observed at moisture contents below 20% is a decrease in extractability of large glutenin aggregates and in hydrophobicity of the acetic acid unextractable protein. At higher moisture contents, also, disulphide bonding and irreversible conformational changes can be observed.