PHYSICAL CHARACTERIZATION OF THE N-TERMINAL DOMAIN OF HIGH-MOLECULAR-WEIGHT GLUTENIN SUBUNIT DX5 FROM WHEAT

The high-molecular-weight (HMW) glutenin proteins from wheal are stora ge proteins. Despite considerable research effort, relatively little i s known about the factors that determine their properties. Their seque nces are homologous and can be divided into three domains which differ considerably in composition. In order to characterise their individua l contributions to the HMW properties, the domains of HMW subunit Dx5 have been cloned. In this paper, the cloning of the N-terminal A-domai n is described together with its production in and purification from E scherichia coli. The protein was insoluble in aqueous solutions under a range of conditions except in the presence of low amounts of sodium dodecyl sulphate. Comparison of the solubility characteristics of the A-domain with those described for HMW Dx5 indicated that the A-domain determined the solubility of HMW Dx5. Changes in circular dichroism an d fluorescence spectra indicated that the structure content of the A-d omain was pH dependent and was highest at low pH. Thermal and urea den aturation studies showed the presence of two independently unfolding p arts in the A-domain; the stability of one decreased with pH, the othe r was less affected by changes in pH. A stretch of 38 residues contain ing the cysteines that are important for the in-vivo crosslinking of H MW proteins is probably responsible for the observed pH dependence. Im plications of the observations for the full-length HMW proteins are di scussed. (C) 1998 Academic Press.