Mutations in the Contact Region between the α and β Subunits of Tryptophan Synthase Alter Subunit Interaction and Intersubunit Communication

Interaction between the α and β subunits of tryptophan synthase leads to mutual stabilization of the active conformations and to coordinated control of the activities of the two subunits. To elucidate the roles of specific residues in the interaction site between the α and β subunits, mutant α and β subunits were constructed, and the effects of mutation on subunit interaction and intersubunit communication were determined. Mutation of either α subunit Asp56 (αD56A) or β subunit Lys167 (βK167T), residues that interact in some crystal structures of the tryptophan synthase α2β2 complex, decreases the ability of the α subunit to activate the β subunit and alters the reaction and substrate specificity of the β subunit. Partial conformational repair is provided by α-glycerol 3-phosphate, a ligand that binds to the α subunit, or by Cs+ or NH4 +, ligands that bind to the β subunit. Mutation of β subunit Arg175 (βR175A), a residue that interacts with α subunit Pro57 in some structures, has much smaller effects on activity but results in a 15-fold increase in the apparent K d for dissociation of the α and β subunits. Replacement of the single tryptophan in the β subunit by phenylalanine (W177F) has only small effects on activity but increases the apparent subunit dissociation constant ∼10-fold. The most important conclusions of this investigation are that interaction between αAsp56 and βLys167 is important for intersubunit communication and that mutual stabilization of the active conformations of the two subunits is impaired by mutation of either residue.