Figure 2.9 Schematic representation of the wild and modified proglycinin A1aB1b. (a) wild type, (b) tetramethionine-inserted proglycinin. White and black areas indicate the conserved and variables regions, respectively. The number of the residues from the N-terminus for the variable regions (I-V) are shown above the alignment. S-S indicates a disulfide bond.

Figure 2.10 Fortification of Met into ^-barrel of the proglycinin A1aB1b. (a) wild type; (b) methi-onine-introduced proglycinin. Backbone and side chains are indicated by bond models.

by the increased hydrophobicity inside the molecules, resulting in the lowering of digestibility. On the other hand, introduction of lysine and arginine residues recognized by trypsin inside the molecule may probably have a large influence on digestibility, because they not only add the site for cleavage, but also destabilize the structure due to the introduction of a charged residue inside the molecule. Most of the salt bridges inside molecules contribute largely to structural stability. Therefore, a substitution of an acidic residue which forms a salt bridge inside the molecule with another amino acid destabilizes a structure while keeping the site for trypsin. In glycinin A1aB1b, a salt bridge between Asp157-Arg161 exists inside the molecule. We replaced Asp157 with Ala (D157A) (54). This mutant, produced in E. coli, was found to fold correctly. Furthermore, we confirmed that D157A formed crystals suitable for X-ray crystallography. The Tm value of D157A was 15°C lower than that of the wild type and its sensitivity to chymotrypsin increased.

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