Abstract Most of the unique properties of waxy wheat have been associated with the lack of amylose, that in turn may affect the mutual interactions between starch and proteins. To address this particular aspect, we carried out molecular, rheological, and calorimetric studies on flours from two waxy wheat lines that were compared with a non-waxy one. Dough thermal properties and water binding capacity were investigated by Differential Scanning Calorimetry (DSC) and by thermogravimetric analysis, respectively. Protein solvation, aggregation, and thiol accessibility were also investigated, together with dough mixing properties and stickiness. Proteins in waxy wheat samples needed more water to complete solvation, likely because of the water-retaining capacity of waxy wheat starch. In waxy wheat dough, water was tightly bound to starch, and DSC studies indicated an increase in gelatinization temperature. Moreover, the low water mobility in waxy wheat resulted in low and retarded gluten hydration and in high stickiness. In samples with the highest stickiness, protein aggregates were stabilized mainly by hydrophobic interactions. Differences between waxy wheat lines may be attributed to a different structural organization of components within each class of biopolymers.